Developmental Biology - Biology 4361

Cell Fate, Potency, and Determination

September 27, October 4, 2005

Fate Mapping
- fate - sum of all structures cell will form in later stages of normal development
- fate map = diagram
- embryonic regions with distinct fate = primordium or rudiment
- cells multiply & move fate maps change
- fate mapping methods
- label cells - watch over time
- fluorescent tags
- label donor cells - transplant
- organisms w/ variable cleavage- different groups of cells can contribute to fate
Clonal Analysis
cell clone - all surviving descendants of single founder cell
- clonal analysis methods:
- fluorescent labeling (resembles fate mapping)
- somatic crossover - generates genetic label - signal not diluted over time
- positioned randomly throughout tissue
- somatic crossover analysis
-outline = cell mixing
- growth kinetics (if daughters = 1/n of organ, then original = n/2)
- e.g. insect epidermis has restriction lines - clones do not cross
= compartments
- compartments filled with several clones = polyclone
- polyclones are unit of size and shape regulation
- polyclones are units of gene regulation
- each compartment has unique combination of active & inactive
selector genes (binary zip code; i.e. sets of genes on or off)
- selector genes - control expression of other genes
Potency of Embryonic Cells
potency - widest range of capabilities of cell
= total of all structures a cell or its descendants can form if placed in the appropriate
environment
- experimental of potency established by operational definitions
- methods:
- isolation - cell (or region) removed from influence of embryo
- in vitro
- heterotopic transplantation - cell or region moved to different region
- cell or region influenced by cells other than normal neighbors
- potency of region always includes its fate (p. 129)

- pluripotent - if cell or region can form more structures than their fate
- totipotent - cell or region can give rise to complete individual
- community effect
- single cells or small number of cells - depart from fate - blend with new region
- larger groups retain original fate
- community effect depends on intracellular communication
Determination
- as development proceeds, range of cell fates diminishes
- when potency of cell restricted to its fate, it is considered determined
- process by which fate & potency becomes identical = determination
- determination methods:
- isolation
- heterotopic transplantation
- clonal analysis
- clone must be restricted to fate
NOTE - clonal analysis used as a negative criterion only; i.e. if
clone differentiates into multiple fates, the cell or tissue was not
determined
- determination proceeds in a stepwise manner
- preceded by bias (preliminary determination)
- methods:
- clonal analysis
- transplantation
- isolation
- cells are born with a bias
- bias confirmed or modified through cell-cell interactions
- determination examples: Drosophila gastrula (see below for notes on gastrulation)
- within prospective thorax and abdomen, dorsal ectoderm forms only epidermis; ventral
ectoderm forms both epidermal and neural cells
- dorsal epidermal analge (DEA) - forms epidermal cells
- ventral neurogenic region (VNR) - forms epidermal and neural
- experiments:
- isolation experiment:
- all DEA cells for epidermis (in accord with fate)
- VNR cells tended to form neurons - (not in accord with fate: mixture expected)
- transplant - switch regions
- DEA to DEA (controls) - mostly epidermal; no neurons
- DEA to VNR - strong community effect; transplanted cell groups formed
epidermis (original fate); single cell transplants - formed neural or mixed
clones
- VNR transplantation - to DEA - single cells neural or epidermal in proportion to
homotopic transplantation; VNR transplanted to other regions = mostly

neural; no epidermal
- results indicates that original vias of VNR cells to form neurons
counteracted more strongly in DEA than in other regions
- bias can be acquired by uneven distributions of cytoplasmic or membrane-bound
activities
- e.g. Drosophila embryo - biases of VNR and DEA traced back to regulatory
protein that accumulates in nuclei of ventral but not dorsal blastoderm
cells
- modification of biases
1. Community effect - exchange among equivalent cells of signals that stabilize the
same determined state for all of them
2. Lateral inhibition (opposite of community effect): equivalent cells compete
with one another to attain a preferred fate; one cell inhibits neighbors from
attaining same fate
3. Embryonic induction - interaction between nonequivalent cells (note Speemans
eye induction experiment)
Further examples of Determination - amphibian neural plate (at gastrulation)
[Gastrulation (gaster - stomach; literally, stomach formation):
NOTE - gastrulation will be covered in detail later
- process of highly coordinated cell and tissue movements whereby cells of blastula
rearranged
- position of blastula cells established during cleavage; gastrulation - cells have new
positions; new neighbors
- multilayered body plan of organism established
- cells that will form endodermal and mesodermal organs brought inside embryo
- cells that form skin and nervous system spread over outside surface
- three germ layers produced (in triploblasts)
ectoderm
mesoderm
endoderm
- stage set for interaction of newly positioned tissues
- gastrulation movements:
- depression forms at blastopore (below equator, near gray crescent)
- vegetal cells buckle into blastula; i.e. move towards blastocoel
- groove (blastopore lip) forms on the dorsal side where cells ingress (move from
epithelium to embryonic cavity)
- groove extends ventrally to form circle
- animal and equatorial cells proliferate and move into blastopore
- animal cells eventually cover yolk; close (blastopore forms anus in
deuterostomes; e.g. vertebrates; mouth in protostomes e.g. arthropods)]
- at late gastrula stage, embryo consists of outer layer - ectoderm and inner layer mesoderm and endoderm

- mesoderm - muscle, etc.

- endoderm - archenteron (embryonic gut)
- ectoderm - epidermis & neural plate
- neural plate - originates from dorsal ectoderm between blastopore and animal
pole
- neural plate is where rudiments of the neural cells are formed
..neural plate rudiment ..fated by not yet determined to become neural plate. p. 135
- i.e. under normal developmental circumstances, neural plate rudiment in late gastrula
would become neural plate; however, could become different tissue if isolated or
transplanted
- determination in gastrula neural plate tested by Spemann:
- transplanted pigmented early gastrula neural plate cells to unpigmented host
- homotopic - formed neural plate
- heterotopic - contributed to epidermis
- conclusion - blastomeres not determined in early gastrula
- repeated with late gastrula neural plate
- homotopic - formed neural tissue
- heterotopic - formed neural tissue
- conclusion - neural plate became determined during gastrulation
Mouse embryonic cell determination
- most, if not all mouse blastomeres up to 8-cell stage are pluripotent
- test by dissociating blastocysts then adding blastomeres back to host blastulas
and following fate
- genetic markers used to follow contribution of blastomeres
- at 16-cell stage, some blastomeres pluripotent, others starting determination process
- mammalian cells from morula through at least 16-cell stage are biased, but not
altogether determined

Properties of the Determined State

- pattern formation - patterns set up from earliest point
- e.g. many eggs have unequal distribution of material - e.g. animal-vegetal
distribution sets up dorsal-ventral pattern
- patterns change with time and developmental age
- pattern formation precedes cell differentiation
- cells determined before reaching terminal differentiated state
- determination assessed by operational criteria
- i.e. tested by isolation, transplantation, etc., not by morphological or
biochemical methods
- e.g. mammalian morula cells: inside and outside morphologically distinct, but
not necessarily determined (i.e. can be interchangeable)

- determination multi-faceted, stepwise process; negative and positive

- loss of potency (potential) = negative or subtractive process
- instruction (i.e. cells need additional instruction (gene activity) in order to follow
alternate pathway) = positive or additive process
- e.g. prospective neural plate would form epidermis in the absence of
further instruction
- instructional mechanisms:
- localized cytoplasmic determinants - cause blastomeres to be determined (or
biased) to form cell lineages or major body regions
- e.g. pole plasm (insects) - form germ cells (pole cells; primordial germ
cells)
- cell-cell communication (intercellular signaling)
- community effect
- induction
- lateral inhibition
- hierarchy of determinative events
- time sequence of cell movement and interaction
- determined state stably passed on during mitosis
- implies genetic and epigenetic controls
- e.g. imaginal discs
- determined state? tested by transplantation
- each imaginal disc is determined
Regulation in Development
- invariant cleavage (e.g. Ceanorhabditis elegans) - precise harmony (i.e. timing and fate)
during development
- variable cleavage (most species) - stepwise approximation to correct imbalances;
development in individual cells can be shifted
- mosaic development - potency map identical to fate map - i.e. all cells determined
(nematode, ascidian)
- regulative development - potency of cells greater than fate; cells not yet determined
(amphibian, sea urchin)
NOTE - in almost all circumstances, development will reach mosaic stage (i.e. virtually all cells
will eventually be determined); how early??
- also, are there regional differences?
- regulation - ability of cell or embryonic region to deviate from its fate (in response to
environmental circumstances)