GENIC CONTROL O F CHROMOSOME LOSS AND NONDISJUNCTION

IN DROSOPHILA MELANOGASTER

RICHARD A. SPIELER2
Department of Zoology, Universiiy of Chicago, Chicago, Illinois
Received September 10, 1%2

HROMOSOME nondisjunction, the failure of the members of a chromosome

pair to separate to the two daughter nuclej, was first described by BRIDGES
(1913,1916). Theoretically, nondisjunction of the X chromosomes in XX females
should result in the production of equal numbers of nullo-X and diplo-X gametes.
Consequently, the two resulting classes of viable exceptional off spring, patro-
clinous sons and matroclinous daughters (presumably XO and XXY, respec-
tively), should be recovered in statistically equal numbers. It was soon realized,
however, that other events partially mimic the consequences of nondisjunction
by giving rise to patroclinous sons but differ in yielding no matroclinous
daughters. In the extensive data of SAPIR(1920) patroclinous sons were found to
be about four times as frequent as matroclinous daughters. A much greater dis-
parity observed in a particular strain of Drosophila mehogaster (SPIELER1961)
led to the investigations described in the present paper.
An excess of patroclinous males would seem to be due to the recovery of more
nullo-X eggs than diplo-X eggs. The unequal recovery could be caused by a
variety of events which will be referred to generically as chromosome loss.
Chromosome loss will be said to occur whenever the ratio of patroclinous males
to matroclinous females is significantly greater than one. This operational
definition must be applied with caution, however, to those cases in which one
might expect important viability differences to result from the relative muta-
tional load of the maternal and paternal sex chromosomes.
Although no attempt will be made to present an exhaustive list of the oc-
currences which may contribute to chromosome loss, a few will be mentioned
briefly. One obvious possibility (which could take place at either meiotic division)
would be separation of the X chromosomes followed by noninclusion of an X in
one daughter nucleus. Or, after nondisjunction has occurred, there may be a
directed orientation of the four meiotic products such that a nullo-X nucleus,
rather than a diplo-X nucleus, more often becomes the egg pronucleus. Also,
heterozygous X-chromosomal inversions produce nullo-X eggs by four-strand
double exchange within the inversions ( STURTEVANT and BEADLE1936) or by
three-strand double exchange, one exchange within the inversion and one outside

1 This investigation was supported in part by Public Health Service Research Grant RG-7428
and by a Traineeship from Public Health Service Training Grant 2G-150.
2 Present address: Dept. of Biology, Illinois Institute of Technology, Chicago, Illinois.

Genetics 48: 73-90 January 1963

74 R. A . SPIELER

and proximal to the inversion (PTASHNE 1960). These eggs are recoverable as
patroclinous males.
An extreme example of chromosome loss is that associated with the claret
mutant stock of Drosophila simulans (STURTEVANT 1929). Fifty percent of sons
and about six percent of daughters are X-chromosome exceptions. Fertilized eggs
from claret females were examined by WALD(1 936). At the end of the first
maturation division the chromosomes lie widely separated from one another at
each pole of an unusually wide spindle. Second division usually fails to occur,
and the cell contains an abnormally large number of nuclei, each containing
less than the haploid complement of chromosomes. There is some evidence of
occasional reincorporation of a chromosome previously eliminated.
The meiotic peculiarities described for claret of D. simulans are seemingly
under genic control. In D. melanogaster the important instances of apparent
genic control of nondisjunction and chromosome loss are those reported for
claret-nondisjunctional (LEWISand GENCARELLA 1952) and for C3G (GOWEN
1933).
In 1960 D R . JOHN L. HUBBYcalled my attention to a mutant strain of D.
nelanogaster in which he had observed a high proportion of exceptional sons.
The strain is maroon-like (ma-1), an eye color mutation which GLASSMAN
(1959) has shown to be located at the proximal end of the X chromosome.
Experiments undertaken to investigate the ma4 strain reveal that the chromo-
some loss found in females of this stock involves a factor at the proximal end of
the X chromosome. The major factor responsible for chromosome loss appears
to be subject to modification by a complex system of sex-linked genes. In
contrast, a stock of rosy, a mutant on the third chromosome, shows a high
incidence of chromosome nondisjunction. The meiotic factor in this instance
appears to reside on the autosomes but affects the disjunctional behavior of the
X chromosomes.
MATERIALS A N D METHODS

Stocks of ma-1 (maroon-like, 1-64.8), ma-l; st, and BX' ma-2 were obtained
from J. L. HUBBY.The stock of ma-1; st (scarlet, 3-44.0) had been synthesized
by crossing ma-1 flies to a stock of st in some numbers, whereas Bx3 ma4
(Beadex3, 1-59.4) had been derived from one male by crossing-over within a
Bx3/ma-l heterozygote formed by crossing the two respective stocks. The stock
of y z ma-lbz (bronzy, an allele of ma-1) was procured through the courtesy of
DR.E. H. GRELLof the Oak Ridge National Laboratory. The stocks involved in
its synthesis are unknown to me. D R . E. GLASSMAN kindly supplied the stock of
ma-lbz.The rosy ( r y , 3-51) and rosy2 stocks came from the California Institute
of Technology.
A standard procedure was devised for deriving those stocks involving crossing-
over between X chromosomes. The initial step (unless otherwise stated) was to
mate flies bearing a given X chromosome to a stock of y 2cu U f. This latter stock
carries the following markers along the X chromosome: yellow2 (yz,0.0) , cross-
veinless (cu, 13.7), vermilion ( U , 33.0), and forked ( f , 56.7). The resulting
 CHROMOSOME LOSS A N D NONDISJUNCTION 75
heterozygous females were then mated to Oregon-R males. In the next generation
a son carrying the desired crossover chromosome was mated to MULLER-5
(Zn~(l)sCls~~,S, scaR,Wa B ) females and subsequently to its own daughters. This
backcross produced some progeny which were homozygous for the crossover
chromosome, and these flies were used to establish the new strain.
Primary nondisjunction and loss of the X chromosomes was detected by cross-
ing females of any given genotype to 'Bar males. Each exceptional (patroclinous)
son was tested for sterility, and many exceptional (matroclinous) daughters
were examined for seconckary nondisjunction. No attempt was made to score
XXX females.
Mass matings were carried out in the preliminary part of the work. Indi-
vidual matings consisted of one female and three Bar males per vial. Some experi-
ments comprised 30 such matings, whereas most of the others were composed of
60 individual matings. Transfers to fresh vials were made every three or four
days. Mass mating results are valid only in a qualitative way, since the resulting
crowding may select against certain genotypes. In addition, crosses made up from
stocks in which nondisjunction occurs with a high frequency will often include
some female parents which are actually XXY in constitution. When using
individual matings one can detect these females by the fertility of their patro-
clinous sons and can exclude their off spring from the data.
In most experiments the offspring reported covered the entire lifespan of the
mothers, which were not more than a few days old when mated.
*
All crosses were maintained in a constant-temperature room at 24 1C on a
yeast-sugar medium with propionic acid to inhibit mold growth.
In computing the frequencies of exceptional gametes produced as a conse-
quence of primary nondisjunction or chromosome loss in females, the following
formulae were applied. Expressions in parentheses represent the number of
progeny of a given type which were recovered.
e(exc.8 8 )
Frequency of nullo-X eggs =
+ + +
(reg. ? ? ) (reg. 8 8 ) 2(exc. ? ? ) 2 (exc. 8 8 ) '
(exc. 8 8 )
which is better expressed as
(reg.? Q)+(exc.?? ) + ( e x c . 8 8 )
, in view of the.
greatly differing viabilities of the regular sons in the various experiments.
Similarly,
(exc. 9 ? )
Frequency of diplo-X eggs =
(reg.? ?)+(exc.? ? ) + ( e x c . 8 8 ) *
The 95 percent confidence limits for the recovered exceptional progeny were
obtained by use of the tables in STEVENS (1942) and in FISHER and YATES(1957).

RESULTS A N D DISCUSSION

Primary nondisjunction and loss in females

a. The maroon-like stock: In the data of BRIDGES(1916) exceptional progeny
were found at a frequency of about one in 1700 (0.06 percent). SAFIR (1920)
76 R. A. SPIELER

tested females of three mutant genotypes and found frequencies of 0.18 percent
0.08 percent, and 0.07 percent. Frequencies for some homozygous stocks used
in the present studies can be found in Table 1. The stocks obviously differ
greatly in their production of exceptions, the cand (claret-nondisjunctional)
stock showing the highest values. Interestingly, the two wild-type stocks produce
more exceptions than do some mutants. Thus there is no correlation between
mutant phenotype and chromosome behavior.
Examination of Table 1 also reveals a varying incidence of chromosome loss.
Although male exceptions almost always outnumber female exceptions in these
experiments, the ratio is only about 2:l in wild type and even lower in some
other cases.
The ma-l stock exhibits a high frequency of chromosome loss. Females produce
nullo-X gametes at a frequency of about 1.2 percent, as compared with 0.1 to
0.3 percent in several non-maroon-like strains. Diplo-X eggs, however, occur
only rarely. In contrast, the stock containing bronzy (an allele of m - 1 ) shows a
very low frequency of nondisjunction or loss, as does the y 2 mcz-Zbzstock. BRANDT-
SNEDECOR tests (SNEDECOR 1956) and contingency table tests revealed a lack of
uniformity between m a 4 females in the proportion of exceptional progeny
produced.
Location of the major factor responsible for chromosome loss: By the standard
crossover technique already described, a crossover on the ma-E X chromosome was
obtained between f (56.7) and ma-1 (64 or 65). In the stock established from this
crossover only the right end of the y Lcu U f ma-l chromosome was derived from
the original ma-2 chromosome. When tested the new stock showed the typical
loss effect (Table 2). Since the autosomes of the ma-Z stock should have been
largely replaced in this strain as a consequence of the outbreeding process
inherent in the synthesis. it can reasonably be concluded that the loss factor
resides on the proximal end of the X chromosome.
That the derivation process itself is not responsible for the effect is shown by
the disjunctional normality of females of the yzcu z? f stock used in the synthesis
(Table 1) and of two strains of y3 cu u f ma-lbz(Table 2) which were derived in
the same way as y' cu Y f mu-l.
Further attempts at localization of the loss factor involved obtaining crossovers
to the right of ma-l. GLASSMAN(1959), BROSSEAU,NICOLETTI, GRELLand
LINDSLEY (1961), and SCHALET (in press) have shown ma-l to be to the left of
su-f (suppressor of forked). Crossover sons were obtained from y' cu U f ma-l
-I-/+ + +
u f su""-f females, and stocks were derived in the usual way. Two
strains of the genotype ++ ++
U f and four strains of its complementary
product y z cu U f m a 4 suW-fwere tested for nondisjunction and loss.
If the loss factor is located on the X chromosome of the original ma-l stock to
+
to the right of su-f. then all strains of the f U f ++ genotype should show the
++ ++
loss effect. Both U f strains behave normally, however, (Table 2) like
the precursor ++ + u f suW-fstock (Table 1) .
Conversely, if the factor is to the left of su-f, at least some cu u f ma-2 suW-f
strains should show the effect. Specifically, if the factor is between f and ma-2
 TABLE 1
Frequencies of nullo-X and diplo'-X eggs from females of certain stocks
Frcq. nullo-X conf. lim. Freq. diplo-X 95% conf. lim.
Expt. Genotype of mothers eggs Percent "7 percent) ems Percent (percent)

W, Z Oregon-R 46/19,641 0.234 0.172- 0.312 26/19,641 0.132 0.086- 0.194

KK Canton-S 24/19,439 0.123 0.079- 0.184 11/19,439 0.057 0.028- 0.101
HH,II,JJ white 5/26,163 0.019 0 . 0 0 6 0.045 1/26,163 0.004 o.Oo0- 0.m1
11 maroon 2/ 7,474 0.027 0.003- 0.097 I / 7,474 0.013 0.000- 0.075
AD U f suw-f 9/ 2,638 0.341 0.156- 0.646 6/ 2,638 0.227 0.083- 0.494
AC y2 cu v f 4/ 2,059 0.194 0.053- 0.497 O/ 2,059 0.000 0.000- 0.179
K scarlet 4/10,786 0.037 0.010- 0.0% 4/10,786 0.037 0.010- 0.095 0
If ClPd 158/ 868 18.203 15.683-20.934 I l l / 868 12.788 10.638-15.195
ma-l (combined 0.013- 0.091 R
E and L) 152/ 12,795 1.188 1.008- 1.391 5/12,795 0.039
E ma-1 22/ 3,342 0.658 0.413- 0.9% 2/ 3,342 0.060 0.007- 0.216'
L ma-l 1301 9,463 1.375 1.150- 1.631 3/ 9,453 0.032 0.007- 0.093
2B
IO* ma-1 28,' 5,886 0.476 0.316- 0.687 IO/ 5,8861 0.170 0.08% 0.312 6
F ma@ 1/ 6,923 0.014 o.oo0- 0.080 1/ 6,923 0.014 0.OOO- 0.080,
6* y2 ma& I/5,959 0.017 0.000- 0.093 o/ 5,959 0.000 0.000- 0.062 5
* Mass matings. a
+ Uncorrected for secondary exceptional females
TABLE 2 4
Nondisjunction and chromosome loss in stocks used to determine the location of the loss factor 2
Freq. nullo-X 95% c o d . lim. Freq. diplo-X 95% cod. lim. $
Expt. Genotype of mothers eggs Percent (percent) eggs Percent (percent)
X,FF,CC y2 cv U f ma-1 39/ 4,928 0.791 0.563-1.080 0/ 4,928 0 0 -0.075 2
y* cv U f ma@ z
DD Strain no. 6 0/ 2,010 0 0 -0.184 o/ 2,010 0 0 -0.184
GG Strain no. 5 2/ 2,418 0.083 0.010-0.299 O/ 2,418 0 0 -0.153
++vf++
xx Strain no. 1 60/13,281 0.452 0.345-0.581 6/13,281 0.046 0.017-0.096
YY Strain no. 2 35/12,608 0.278 0.193-0.386 15/12,608 0.119 0.0674.196
ys cu U f ma4 sulv-f
AF Strain no. 1 14,' 1,877 0.746 0.408-1.248 1/ 1,877 0.053 0.001-0.295
zz Strain no. 3 O/ 471 0 0 -0.783 O/ 471 0 0 -0.783
AB Strain no. 4 3/ 1,182 0.254 0.052-0.740 O/ 1,182 0 0 -0.312
AE Strain no. 5 1/ 2,928 0.034 0.001-0.190 o/ 2,= 0 0 -0.126 -I
w
78 R. A. SPIELER

(or is identical with ma-1),then all y2 cu u f ma4 suW-f strains should exhibit
chromosome loss. But if the factor is between ma-1 and SU-f, then only those
strains in which the particular crossover is to the right of the factor will show the
loss effect. Of the four, only strain Number 1 (Table 2) seems to duplicate the
chromosome loss of the precursor y 2 cu u f ma-1 stock. The data are consistent
with the hypothesis that the loss factor has its locus between ma4 and su-f. The
ma-1 allele per se is not sufficient to produce the loss effect. Still, on the basis of
present information, one cannot rule out the possibility that the presence of ma-1
is necessary for the effect.
Nature of the phenomenon: A high yield of exceptional males, such as ob-
served in females of the ma-1stock, can result from several different phenomena.
Undetected inversions in some of the X chromosomes, for example, could result
in the production of patroclinous sons. This possibility can be ruled out on the
basis of Experiments 7 and 8 (Table 3), which tested females bearing one ma4
X chromosome and one X chromosome from Oregon-R. Inversion heterozygotes
seem to be absent, since no patroclinous males are produced.
If pairing difficulties exist, for whatever reasons, between the Xs carrying
the loss factor, then the presence of a heterozygous autosomal inversion might
lead to increased nonhomologous pairing between the Xs and autosomes and so
to the production of more exceptions (COOPER,ZIMMERING and KRIVSHENKO
1955 and ROBERTS unpublished). A stock was constructed (using multiple-
inversion chromosomes) which contained the first, second, and third chromo-
somes of the ma4 stock. Some individuals, however, carried a C y or a Ubx
chromosome (or both) in place of one second or third chromosome from the
ma-1 stock. Females of three genotypes were tested (+ indicates an autosome
from the ma-1 stock) :
ma-1 C y Ubx
1) -.- -(Experiment UD)
ma-17 + +
;
ma-l
2) -. C y
-. - $- (Experiment AM)
ma-1 + +

3) -*
ma-1 + -
-* +
+ (Experimentm)
+

The results (Table 3 ) show no significant difference between genotypes in

the yield of X-chromosome exceptions. The absence in experiment AM of any
great increase in exceptions argues against nonhomologous pairing as an im-
portant component of the chromosome loss effect. Nonhomologous pairing should
result in equal classes of nullo-X and diplo-X eggs. Even if there were a directed
meiotic orientation for preferential recovery of nullo-X eggs, the frequency of
patroclinous males, at least, should increase in Experiment AM. THOMPSON
(1962) studied pairing in ma-1 by using a 3;4 translocation and also reached the
conclusion that failure of pairing is not the reason for the chromosome loss
effect.
Although nonhomologous pairing can be discarded as a causative factor, there
remains a variety of possible reasons for production of nullo-X eggs in females
 TABLE 3
Production of exceptions in stocks used to determine the nature of the chromosome loss phenonzenon

Freq. nullo-X 95% conf. lim. Vreq. diplo-X 95% conf. lim.
Expt. Genotype of mothers eggs Percent (percent) eggs Percent (percent)

8* mad/+ 0/5,291 0 0 -0.070 3/5,291+ 0.057 0.012-0.166

7* mad/+ ;st/+ 1/1,253 0.080 0.002-0.445 1/1,253+ 0.080 0.00.%0.445
ma-1 Cy U b z
UD __ .._ 13/3,323 0.391 0.232-0.668 ...... .... ..........
ma-1 -I jt- 89
ma-1 c y .+ 0
AM -. - 52/9,977 0.521 0.3!30-0.683 3/9,977 0.030 0.006-0.088
mad ?t;
+ 8
ma-1 + + 8
AN . 17/5,121 0.332 0.193-0.531 0/5,121 0 0 -0.072
ma-1
-. _ . _
+ + Er
matings. $
Uncorrected for secondary exceptional females.
+* Mass ul

TABLE 4
%
U

Modification phenomena in ma-1; st and in some Bx3 ma-1 stocks z

4
Freq. nullo-X 95% conf. lim. Freq, diplo X 95% c o d . lim. E!
V)
Expt Genotype of mothers eggs Percent (percent) eggs Percent (percent)

C ma-1; st 56/2,854 1.962 1.4862.541 28/2,854 0.981 0.653-1.415

2
4* ma-2; st 81/7,416 1.092 0.868-1.356 27/7,41w 0.364 0.2404.530 3
El% ma-2; st (new stock) 0/4,632 0 0 -0.080 0/4,632 0 0 -0.080 8
7
D Bx3 ma4 4/5,662 0.071 0.019-0.181 2/5,662 0.035 0.004-0.12.8
3* Bx3 ma-1 2/5;3 11 0.038 0.005-0.136 0/5,3 11 0 0 -0.069
ma-Iex Bx3 ma-l
AA Strain no. 1 35/4,389 0.797 0.5561.107 34/4,389 0.775 0.537-1.0.81
BB Strain no. 2 89/4,838 1.840 1.480-2.259 79/4,838 1.633 1.295-2.031
matings.
Uncorrected for secondary exceptional females.
+* Mass U
rD
80 R . 4 . SPIELER

of the ma-1 stock. Further progress in the analysis of the phenomenon depends on
devising methods for distinguishing between these possibilities.
A rather puzzling point was the discovery during the course of these experi-
ments of three apparent attached-X daughters. The mother of one was ma-1; st
(Experiment C) and was XX, as shown by the sterility of all six of her excep-
tioaal sons. She produced four other exceptional daughters, three of which were
tested and which behaved genetically as ordinary XXY females. The mother of
the other two was ma-1; Cy; Ubx (Experiment UD). She produced three ex-
ceptional daughters, one of which was not saved. The other two behaved
genetically as attached-X females, producing all matroclinous daughters and
patroclinous sons. One daughter was simply ma-l/ma-1, whereas the other was
ma-l/ma-1; Ubx/+. Contamination can be ruled out as an explanation, since no
ma-1 compound-X chromosomes were maintained in the laboratory.
The basis for these two independent origins of compound-Xs is unknown, but
it is conceivable that at least some ma-Z Xs possess an extra arm, crossing-over
then resulting in occasional attachment of the Xs. Cytological examination of
larval neuroblasts in the m a 4 stock failed to reveal any visible extra arm. These
cases are very suggestive, however, of some structural abnormality in the centro-
meric heterochromatin of some or all of the ma-l X chromosomes.
Modification of the phenomenon: The meiotic properties of the m a 4 stock are
subject to both qualitative and quantitative changes when the genetic background
is altered. A remarkably higher frequency of exceptions is found in a mal-1; st
stock synthesized by HUBBY (Table 4). Frequencies of nullo-X and diplo-X eggs
are increased to about 2 percent and 1 percent, respectively, as if the occurrence
of true nondisjunction at about 1 percent had been superimposed on the 1 percent
of chromosome loss found in the original ma-1 stock. Whether this apparent
introduction of true nondisjunction can be attributed to sex-linked or autosomal
factors (or both) has not been determined. In the synthesis of the strain there
was opportunity for crossing-over on the Xs as well as introduction of new
autosomes from the st stock.
A new synthesis, using the same ma-l stock but another stock of st, gave a strain
of ma-Z; st which yielded no exceptions whatsoever (experiment EE) . The same
stock of st by itself produced few exceptions (Table 1). Suppressors of the loss
factor may have been introduced from the st stock.
A stock showing a very low incidence of exceptions, Bx3 ma-1, was more
thoroughly investigated. The stocks in Experiments AA and BB came respectively
from two sons of BxS ma-Z/Ore-R mothers. The Bx3 marker having been elimi-
nated by crossing-over, the two new ma-1 stocks were made up by the usual
method. When tested both stocks gave high frequencies oE exceptions (Table 4).
Interestingly, the sex ratio of exceptions is 1: 1. The results, therefore, must
reflect the occurrence of true nondisjunction rather than chromosome loss. In
any event, the alteration of the genome of the Bx3 ma-1 stock, with respect both
to the X chromosomes and the autosomes, results in the occurrence of meiotic
abnormalities. It would seem that the low frequencies observed in the Bx3 ma-]
stock can be attributed to suppression of the loss factor and not to its absence.
 C H R O M O S O M E LOSS A N D N O N D I S J U N C T I O N 81
Are these suppressors sex-linked, autosomal, or both? In Experiments AA and
BB all the X chromosomes within a strain were identical by virtue of the
derivation process. Any heterogeneity between mothers in the production of
exceptional offspring should be due to the assortment of autosomal loci. Both
experiments when analyzed by tests of homogeneity do implicate the autosomes.
But the quantitative and even the qualitative role of the autosomes in modifying
the loss factor is not yet clear.
T o study the influence of the X chromosome on suppression, Crossover stocks
I I1 I11 IV v
were synthesized following the standard scheme. That is, yz cu U f + +/+
4-++ Bx9 mu-1 females were made up, and single crossovers were procured in
each of the five regions and made into stocks. Three different crossover strains
from each region were then tested. Within each strain all X chromosomes were
identical, but the autosomes were a mixture derived from several stocks. Rela-
tively few autosomes from the BxS m-1stock should remain in the new strains.
This plan has the effect of progressively including in the X chromosomes of the
new strains additional segments of the original BxS ma-1 chromosome. In the
following discussion I f will represent the major factor responsible for chromo-
some loss.
Single crossovers in Region V: All three strains representing crossovers
between Bx and mad, show the loss effect (Table 5 ) . The chromosome can be
represented as yzcu U f Bx+ ma4 I f .
Single crossovers in Region IV: All crossover strains from Region IV (between
f and B x ) also show chromosome loss: p cu U f BxS ma-1 I f .
Single crossovers in Region 111: Again all strains exhibit uniform frequencies,
but in this case the incidence of nullo-X eggs is only 0.2-0.3 percent. It appears as
if the Bx3 ma4 X chromosome contains a suppressor (m,) in the region between
U and f . All three strains would represent crossovers to the left of m,: ye cu u m,
ifBx3 ma-1 I f .
Single crossovers in Region 11: When single crossovers take place between cu
and U the chromosome loss effect is regained. Since suppressor m, is present, it is
necessary to postulate the existence of a second modifier, m2,which suppresses
the action of factor m,. All three crossovers in this region must have been to the
left of m2:y 2 cu m, U + m, f + Bx3 ma-Z I f . Since these stocks show chromosome
loss, whereas the full Bx3ma-Zchromosome exhibited little loss, one can predict
the existence of still another factor, somewhere to the left of m,, which should
suppress the action of mz so that there will again be suppression of the loss effect.
Single crossovers in Region I: Two strains (Numbers 1 and 4) produce excep-
tions at the high level characteristic of the loss effect. In the other strain (Number
2), however, there is very little loss. These results tend to verify the existence of
another modifier (m,),located between y and cu.
The first two strains must represent crossovers to the right of m3and the third
strain to the left of that factor.
Strains 1 and 4: y s cuf m, U + m, f + BxSma-Z If
Strain 2: yz m, cu+ mpU + m, f + Bx9 ma-1I f
 TABLE 5
Modifications of the loss effect by the euchromatic regions of the Bx3 ma-1 chromosome
Regions I I 1 III IV V
Crossovers derived from y2 + + females
+ + f -k Bz3 ma-l
Freq. nullo-X 95% conf. Inn. Freq. diplo-X 95% c o d . lim.
Expt. Genotype of mothers eggs Percent (percent) ems Percent (percent)
_ _ _ _ _ ______
SCO, Region V
y8 cu U f + ma-l
Combined 73/ 6,860 1.064 0.835-1.336 I/ 6,860 0.015 0.000-0.081
00 Strain no. 3 38/ 2,835 1.341) 0.950-1.835 I/ 2,835 0.035 0.001-0.196
PP Strain no. 6 15, 1,868 0.803 0.450-1.322 0/ 1,868 0 0 -0.198
QQ Strain no. 1 20/ 2,157 0.927 0.568-1.428 0/ 2,157 0 0 -0.171
SCO, Region IV
ys cu U f Bxj ma-1
0
Combined 174/12,411 1.402 1.203-1.624 18/12,411 0.145 0.0860.229
LL Strain no. 4 46/ 5,439 0.846 0.620-1.126 4/ 5.439 0.074 O.OuFo.188 ?
MM Strain no. 2 38/ 3,792 1.002 0.710-1.373 3/ 3,792 0.079 0.0160.231 v,

NN Strain no. 5 90, 3,180 2.830 2.282-3.468 11, 3,180 0.346 0.173-0.618 $
SCO, Region I11 FF
y2 cu U + Bxs ma-1
Combined 19, 7,385 0.257 0.156-0.401 4/ 7,385 0.054 0.0815-0.139
RR Strain no. 1 5/ 2,400 0.208 0.068-0.485 3/ 2 , m 0.125 0.026-0.365
uu Strain no. 2 6/ 2,049 0.293 0.107-0.636 1/ 2,049 0.049 0 001-0.272
W Strain no. 6 8/ 2,936 0.272 0.1 18-0.536 O/ 2,936 0 0 -0.126
SCO, Region I1
yz cu ++ B r 3 ma-l
Combined 71, 7,369 0.9863 0.753-1.214 3/ 7,369 0.041 0.008-0.119
ss Strain no. 5 25/ 3,450 0.725 0.470-1.068 O/ 3,450 0 0 -0.107
TT Strain no. 3 27/ 2,293 1.177 0.778-1.708 I/ 2,293 0.044 0.001-0.2A.2
ww Strain no. 4 19/ 1,626 1.I 69 0.703-1.821 2/ 1,626 0.123 0.015-0.443
SCO, Region I
yg + f f Brs ma-1
AR Strain no. 4 61/ 8,013 0.761 0.583-0.977 8/ 8,013 0.100 0.04-3-0.197
AS Strain no. 1 64/ 7,025 0.911 0.702-1.162 18/ 7,025 0.256 0.152-0.405
AT Strain no. 2 12/ 9,029 0.133 0.069-0.232 6/ 9,029 0.066 0.0244.145
 CHROMOSOME LOSS A N D NONDISJUNCTION 83
The proposed constitution of the original Bx, ma-2 X chromosome, therefore,
is y+ mscvf m, U + ml f + BxS ma-2If.
The generally similar behavior of the strains Within each genotype in these
crossover studies indicates that autosomal modifiers are probably not affecting
the results under discussion. The constitution of the euchromatic region of the x
chromosome itself plays the determining part under these circumstances in the
expression of the loss effect. These results lead one to postulate the existence of a
suppressor (m,) of a suppressor (m,) of a suppressor (m,) of the major loss
factor.
Interactions in heterozygotes: Tests of mu-Z/+ and ma-l/+; st/+ females show
that the meiotic factor behaves as a recessive. But when different chromosomes of
the general ma-1 system are put together in various heterozygous combinations,
intermediate frequencies of exceptions often result. Certain combinations, such
as ma-Z/yzma-lbzand even BxS ma-Z/yz ma-lbz, yield nullo-X eggs at a frequency
of about 0.6 percent; the recovery of exceptional daughters is normal. In the case
of heterozygotes involving Bxs ma-1, suppressors are then also made heterozy-
gous, explaining the interaction effect, at least in part.
Bronzyinteractions must involve different suppressors or a modified loss factor.
Crossing out most of the original ma-lbzchromosome and simultaneously out-
breeding with respect to the autosomes of the bronzy stock (Experiments DD
and GG, Table 2) leads to no increase in exceptions. This would seem to rule
out suppressors not closely linked to the bronzy locus.
b. T h e rosy stock: Flies from another stock, rosy, exhibit a high frequency of
X-chromosome nondisjunction (Table 6 ) . In this stock both exceptional males
and exceptional females are encountered at a high frequency. In Experiment N
nullo-X eggs exceed diplo-X by a factor of more than two, but the results of
Experiment J would indicate a 1:1 ratio. There are also sizable differences
between the two experiments in the figures for patroclinous males. All these
discrepancies may be attributable to errors of sampling, as can be seen from the
wide confidence intervals. The mothers in each experiment are homogeneous in
their production of exceptions, BRANDT-SNEDECOR homogeneity tests giving a P
value of more than .10 for Experiment J and more than .20 for Experiment N.
A stock of another allele, rosy2, does not show an abnormal incidence of
exceptional offspring, nor do r y / r p females.
The question now arises as to whether the nondisjunction of the X chromo-
somes in the rosy stock is due to factors on the X chromosome or to autosomal
factors. The ma-P; r y females tested in Experiment AL are from a strain having
the Xs from the ma-lbzstock and the second and third chromosomes from the
rosy stock. Although the bronzy Xs ordinarily show normal disjunction, when
combined with rosy autosomes they give frequencies characteristic of the rosy
stock.
The opposite approach is to obtain a stock having the X chromosomes from the
rosy stock but without the presence of rosy itself. Mating rosy males to MULLER-5
females and then pair mating the F2resulted in several strains in which rosy was
altogether absent. One strain was tested (Experiment AZ), and no exceptions
 00
P

TABLE 6

Production of exceptions inuoluing the rosy system

Freq. nullo-X 95%, conf. lim. Freq diplo-X 95% conf. lm.
Expt. Genotype of mothers eggs Iercent (percent) eggs Percent (percent) 0

r y (combined J and N) 44/ 3,294 1.336 0.972-1.790 29, 3,294 0.880 0.590-1.262 ?
J rY 20/ 2,018 0.991 0.607-1.527 19, 2,018 0.942 0.568-1.467
N Y 24/ 1,276 1.881 1.2.09-2.786 10/ 1,276 0.784 0.377-1.437 M
V rY 5/ 7,499 0.067 0.022-0.155 3/ 7,499 0.040 0.008-0.1 17
5* rY2 1/ 6,463 0.015 O.OW0.086 I/ 6,463 0.015 0.000-0.086 E
S rY/rY2 U)/ 12,729 0.157 0.09&0.243 12/12,729 0.094 0.049-0.165
AL ma-lbz; r y 37, 3,870 0.956 0.674-1.315 20/ 3,870 0.517 0.3160.797
AZ Xs from r y stock O/ 5,130 0 0 -0.072 O/ 5,130 0 0 -0.072
Rlasc niatings.
 CHROMOSOME LOSS AND N O N D I S J U N C T I O N 85
were found. Clearly the X chromosomes of themselves cannot account for the
nondisjunction which characterizes the rosy stock.
Thus the rosy genetic system provides an example of a genically controlled
interchromosomal effect.

Primary nondisjunction and loss in males

In the course of the present work several stocks were synthesized to study
chromosome nondisjunction and loss occurring in the XY male. Strains were
derived whose X, second, and third chromosomes were those of the respective
parent stock, but in which the males carried a Bar-Stone Y chromosome instead
of the unmarked Y of the parent stock. By crossing males of these new stocks to
white females, it was possible to score exceptions arising from nondisjunction or
loss in either parent. The production of equational nondisjunctional sperms, how-
ever, would not be detected. I n order to recover a large number of gametes from
each male, an individual mating in these experiments consisted of one male
crossed to three white females. Thirty individual matings composed each
experiment.
The frequencies of exceptional sperms are determined from the following
formulae, where superscript M represents a maternal chromosome and super-
script P a parental chromosome, and where
(reg. ? P ) = number of (XMXP)flies recovered
(reg. 8 8) = number of (XnrYp)flies recovered
(exc. P P ) = number of ( XMXpYp)flies recovered
(exc. 8 8 ) = number of (XOP) flies recovered
(exc. 8 8 )
Frequency of nullo-I sperms =
(reg. ? ?)+(reg. 8 8)+(exc. ? ?)+(exc. 8 8 )
(exc. 8 8 )
which is better expressed as
2 (reg. P P)+(exc. P Q)+(exc. 8 8 )

in order to compensate for the lowered viability of regular males as compared

with regular females. Similarly,
(exc. 0 Q)
Frequency of diplo-I sperms =
2 (reg. ? ?)+(exc. ?)+(ext. 8 8 ) *
A few maternal exceptions (XpOMand XXMYP) were also recovered (Table 1).
It is apparent that nondisjunction and loss of sex chromosomes in the male
takes place at rather low frequencies in all three stocks (Table 7 ) . Although the
proportion of diplo-I sperms is constant in all stocks, there is a real and surprising
difference in the production of nullo-I sperms. In the ma-Z stock the two types
of exceptional sperms are equally frequent. There is no excessive loss in ma-l
males, unlike the situation in ma-Z females.
With Oregon-R and mad ;st males the nullo-I sperms recovered are three or
four times more frequent than diplo-I sperms. There would seem to be a sizable
86 R. A. SPIELER

loss component. A certain proportion of this, however, could be the result of

equational nondisjunction rather than chromosome loss. There is no direct way
to measure the equational contribution in this scheme, since the complementary
equational classes (XMXpXpand XYPYP zygotes) are not easily recognizable.
KELSALL(1961) reports the approximate frequency of XYPYP sons as 0.004
percent. On this basis nullo-I equational sperms should occur about 0.008 percent
of the time; thus these cannot account for more than a small part of the observed
excess of nullo-I exceptions.
Some other possible explanations for the commonly observed excess of nullo-I
exceptions are (1 ) loss of one of the sex chromosomes from a potential diplo-I
spermatocyte, yielding a regular haplo-I sperm, (2) loss of an X or a Y from a
regular haplo-I spermatocyte, or (3) chromosome loss during first cleavage.
RUSSELL (1961) believes the last event to be a major factor in the excess of XMOP
exceptions in mice. These exceptions occur at a frequency of 0.76 percent in her
experiments, a considerably higher frequency than that found in Drosophila
melanogaster.
KELSALLS preliminary data indicate a frequency of about 0.01 percent for
diplo-I sperms, but his computational methods are not explained. FROST (1961),
using a sc*.Y chromosome, obtained frequencies (adjusted to be comparable to
those computed here) of 0.059 percent for nullo-I and 0.033 percent for diplo-I
sperms.

Autosomal and somatic nondisjunction and loss

No attempt was made to obtain evidence of loss of the large autosomes in ma-Z
females. If autosomal loss occurred at about the same frequency as loss of the Xs,
the consequent lethality would not be detected by the relatively insensitive egg-
count method.
Haplo-IV individuals are not encountered in an abnormally high proportion
among the progeny of ma-1 or r y mothers. THOMPSON (1962), using a y2 ma4
stock, recovered nullo-X eggs at a frequency of 1.22 percent but found no haplo-
IV flies among 4,617 offspring.
Somatic nondisjunction or loss probably take place only rarely in the ma4 or
r y stocks, judging from the infrequent occurrence of gynandromorphs (one
major gynandromorph and 12,637regular daughters from ma-1 and none in 3,220
from r y ) .

Secondary nondisjunction
BRIDGES (1916) presented very extensive data on segregation of the sex chromo-
somes in XXY females. Recovery of XXMYP and XpYMindividuals (so-called
secondary exceptions) was about 4.3 percent of each sex in a total of 56,474 off-
spring. Frequencies were extremely variable, however, ranging from less than
1 percent to as high as 42 percent. This wide variability has been reported since
by many authors, and the cause has been laid variously to the constitution of the
Xs, the Y, the autosomes, or all these elements together.
 TABLE 7
Nondisjunction and chromosome loss in males

VI-. nullu-I 95% conf. lini. Freq. diplo-l 95% cunf. lim.
Expt. Genotype of mothers swims Peirent (percent) Sperms Percent (percent)
~

HH ma-I/YB" 5/18,788 0.027 0.009-0.062 5/18,788 0.027 0.004-0.062

I1 + / Y B 8 (Ore-R) 19/18,337 0.104 0.062-0.162 6/18,337 0.033 0.012-0.071
JJ ma-I/YBs;st 16/15,194 0.105 0.060-0.171 4/15,194 0.05% 0.007-0.067

TABLE a
Secondary nondisjunction in primary exceptional females v
0,
0
%
Mothers of Primary exceptions Progeny M
primary exceptions
Nu indlv hxc Reg Exc
% (XX)-(Y) Reg 5
Expt Genotype Sex chroniosnmes tested VI
v,
? ?_ _ _? ?_ _ _ _
d d_ _ _dd
_ _ _ se@-.-
W, Z Oregon-R +/+/Y 16 273 7,926 273 8,089 6.M
KK Canton S +/+/Y 8 89 4,509 75 4,615 3.22
2
tl

11 maroon +/sly 1 9 839 8 931 1.87 2

K scarlet +/+/Y 4 27 1,403 23 1,m 3.17 2
J rY +/+/y 15 67 6,315 42 5,454 1.31
v rYz +/+/Y 2 7 692 5 725 1.42 d
S ry/ry2 +/+/Y 11 77 4,714 76 4,557 3.12 7i
AL ma-Pa; r y ma-P/ma-Pt//Y 6 25 618 21 555 6.36 =i
sz
E ma-I ma-l/ma-l/Y 2 6 271 5 224 3.56
L ma-1 ma-l/ma-l/Y 2 3 216 1 174 0.92
F ma-lba ma-l"/ma- P / Y 5 17 887 29 736 6.14
C ma-1;st ma-l/ma 1/Y 26 149 3,728 144 3,096 7.17
AM ma-1;C y ma-l/ma-l/Y 1 2 96 2 81 4.00
D Bx3 ma-l Bx3 ma-l/Bx3 ma-l/Y 2 12 473 11 318 4.44
AA ma4 ex Bx3 ma-1 ma-l/ma-l/Y 10 39 978 52 706 9.61
BB ma-1ex Bx3 ma-1 ma-l/ma-l/Y 13 75 973 91 770 15.76 or,
U
88 R . A . SPIELER

The frequency of (XX)- (U) segregation was computed using the formula
below, where

(reg. 0 0 ) = number of (XRrXp)and ( XpXMYM) flies recovered

(reg. 8 8 ) = number of ( XMYP)and (XbfYMYp) flies recovered
(exc. 0 0 ) = number of (XnlX&rYp) flies recovered
(exc. 8 8 ) = number of (XPYh1)flies recovered

Percentage of (XX) - (U) segr. =

100 [ 2(exc. 0 ? ) + 2 ( e x c . d 8 )
(reg. ? Q ) + ( r e g . 8 8 ) + 2 ( e x c . ? ? ) + 2 ( e x c . 8 8 ) 1.
Since regular males and regular females should be produced in equal numbers,
and since exceptional males should be as frequent as exceptional females, the
expression above can be written as

100 [2(reg. 0(exc.

4
?)+4(exc.
88)
88) 1
2 (exc. 8 8 )

This form has the advantage that experiments can readily be compared, since
in most experiments the regular females are heterozygous Bar and the exceptional
males hemizygous Bar.
Inspection of Table 8 reveals that, allowing for viability differences, the sex
ratio of exceptions is approximately equal to one, as would be expected. It should
be noted that if nullo-I eggs occur these would be confounded with Y-bearing
eggs, since exceptional males (XpYx and possibly XpOM)were not tested for
fertility.
The highest frequency of secondary nondisjunction in these experiments is
about 16 percent, but most values are far below that level. Frequencies also vary
greatly between females of the same genotype.
The Oregon-R frequency is just about twice that found with Canton-S. It will
be recalled that Oregon-R XX females produce twice as many nullo-X eggs as
do Canton-S XX females.
Another interesting difference is the considerably higher frequency in ma-l/
ma-Z/Y females when these come from ma-Z; st mothers rather than from m a 4
mothers. Surprisingly enough, however, ma-lbaX's (from experiments F and AL)
also show a relatively high value.
The very high incidence of secondary nondisjunction in XXY females from
experiments AA and BB can scarcely be attributed to structurally-determined
pairing difficulties between the X chromosomes, since within each stock all the
X chromosomes are supposedly identical. One would expect optimal pairing to
occur if structural identity of the X chromosomes were the only necessary
condition.
 CHROMOSOME LOSS AND N O N D I S J U N C T I O N 89
SUMMARY

Three important instances of supposed genic control of chromosome nondis-

junction and loss have previously been reported for Drosophila. In this paper
two more cases in D. melanogaster are described.
One system (discovered in a stock of maroon-like) involves a factor located at
the proximal end of the X chromosome which when homozygous in females
produces a high incidence of meiotic loss of the X chromosome. The action of the
loss factor appears to be subject to modification by a complex system of sex-
linked genes which is thought to include a suppressor of a suppressor of a sup-
pressor of the loss factor. In other genetic backgrounds the meiotic properties
are variously altered. In one stock frequent chromosome nondisjunction seems
to be added to the chromosome loss effect. All these phenomena are operative
only in the female; males show no meiotic abnormalities.
The second case (discovered in a stock of rosy) has not yet been analyzed so
extensively, but it seems to differ from the first in at least two interesting respects.
Most of the exceptional progeny result from nondisjunction rather than from
chromosome loss. Secondly, the meiotic factor appears to reside on the autosomes,
but it affects the disjunctional behavior of the X chromosomes. This is, there-
fore, an example of a genically controlled interchromosomal effect.

ACKNOWLEDGMENTS

I should like to express my thanks to DR.WILLIAMK. BAKERfor his kind aid

and interest in this project and to DR. JOHNL. HUBBY
for suggesting the prob-
lem. Grateful acknowledgment is also made to DRS.JANICE B. SPOFFORD and
EILEENS. GERSHfor their help in various ways.

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90 R. A . SPIELER

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