1. Cellular Mechanisms of Development
Chapter 19

2. Process of development
Process of systematic gene-directed changes throughout an organism’s life cycle
4 subprocesses:
Cell division
Differentiation
Pattern formation
Morphogenesis

3. Cell Division Very first process that must occur during embryogenesis
After fertilization, the diploid zygote undergoes a period of rapid mitotic divisions
In animals, controlled by cyclins and cyclin-dependent kinases (Cdks)
Exert control over checkpoints in the cycle of mitosis

4. Cleavage
In animal embryos, period of rapid cell division following fertilization
Blastomeres – Enormous mass of the zygote is subdivided into a larger and larger number of smaller and smaller cells
Not accompanied by any increase in the overall size of the embryo
G1 and G2 phases of cell cycle short or eliminated

6. Caenorhabditis elegans
One of the most completely described models of development
Adult worm consists of 959 somatic cells
Transparent, so cell division can be followed
Researchers have mapped out the lineage of all cells derived from the fertilized egg
Fate of each cell is the same in every C. elegans individual

7. Cell differentiation
Human body contains 210 major types of differentiated cells
Cell determination – molecular decision to become a particular type of cell
Cells become determined prior to differentiation
Standard test for determination is to move cell

9. Determination has a time course
Depends on a series of intrinsic or extrinsic, or both events
Early tissue will turn into something else but not after a certain time
Determination often takes place in stages
Cell first becomes partially committed
Acquires positional labels that reflect its location in the embryo
Tissue determined as leg but not what part of leg

10. Molecular basis of determination
Cells initiate developmental changes by using transcription factors to change patterns of gene expression
Once the initial “switch” is thrown, the cell is fully committed to its future developmental path
Cells become committed via
Differential inheritance of cytoplasmic determinants
Cell–cell interactions

11. Differential inheritance of cytoplasmic determinants
Tunicates are marine invertebrates
Swimming tadpolelike larval stage
Muscles that move the tail develop on either side of the notochord
Colored pigment granules asymmetrically localize to tail muscle cell progenitors

13. Female parent provides egg with mRNA encoded by macho-1 gene
Experimentally shifting colored pigment granules causes other cells to become muscle cells
Female parent provides egg with mRNA encoded by macho-1 gene
Gene product has been shown to be a transcription factor that can activate the expression of several muscle-specific genes

14. Induction Change in cell fate due to interaction with an adjacent cell
Demonstrate the importance of cell–cell interactions in development by separating the cells of an early frog embryo and allowing them to develop independently
Ectoderm develops from cells of the animal-pole
Endoderm develops from cells of the vegetal-pole
No mesoderm develops unless animal-pole cells and vegetal-pole cells are placed next to each other

15. Another example of inductive cell interactions is the formation of the notochord and mesenchyme in tunicate embryos
Muscle, notochord, and mesenchyme all arise from mesodermal cells

16. Copyright © The McGraw-Hill Companies, Inc
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
FGF signaling
Anterior
Posterior
32-Cell Stage
64-Cell Stage b. c.
Prospective mesodermal cells receive signals from the underlying endodermal precursor cells that lead to the formation of notochord and mesenchyme

17. Stem cells
Cells that are capable of continued division, but can also give rise to differentiated cells
Degree of determination
Totipotent – cell that can give rise to any tissue in an organism (embryo and extraembryonic membranes)
Pluripotent – give rise to all cells in the adult organism’s body
Multipotent – give rise to limited number of cells
Unipotent – give rise to only a single cell type

18. Embryonic stem cells (ES cells)
Form of pluripotent stem cells
Made from mammalian blastocysts
ES cells isolated from inner cell mass and grown in culture
In mice, have been shown to develop into any type of cell in the tissues of the adult
Cannot develop into extraembryonic membranes
1998 – first human ES cells
Great promise and controversy

20. Colony of undifferentiated human embryonic stem cells surrounded by fibroblasts (elongated cells) that serve as a “feeder layer”

21. Nuclear reprogramming
Experiments carried out in the 1950s showed that single cells from fully differentiated tissue of an adult plant could develop into entire, mature plants
Cells of an early cleavage stage mammalian embryo are also totipotent
Natural twinning
Producing multiples artificially for commercial agricultural lines of cattle

22. Early experiments showed nuclei could be transplanted between cells
Cells do not appear to undergo any truly irreversible changes, such as loss of genes
More differentiated the cell type, the less successful the nucleus in directing development when transplanted
Nuclear reprogramming – nucleus from a differentiated cell undergoes epigenetic changes that must be reversed to allow the nucleus to direct development

23. Early amphibian work showed that adult nuclei have remarkable developmental potential, but cannot be reprogrammed to be totipotent
Nuclear transfer in mammals did not result in reproducible production of cloned animals
Did lead to discovery of imprinting

24. 1984 – sheep was cloned using the nucleus from a cell of an early embryo
1996 – Dolly, the first clone generated from a fully differentiated animal cell
Used somatic cell nuclear transfer (SCNT)
Dolly matured into fertile adult
Established beyond all dispute that determination in animals is reversible

25. Reproductive cloning
Uses SCNT to create animal genetically identical to another
Efficiency is quite low and other problems
Only 3–5% of adult nuclei transferred to donor eggs result in live births
Due to lack of genomic imprinting
Normal mammalian development depends on precise genomic imprinting
Organization of chromatin in adult and embryo very different

26. Much work has been put into trying to find ways to reprogram adult cells to become pluripotent cells without the use of embryos
Different lines of inquiry showed that reprogramming of somatic nuclei was possible
2006 – genes for 4 different transcription factors introduced into fibroblast cells in culture
Named induced pluripotent stem cells (iPS cells)
Appear to be similar to ES cells in terms of developmental potential, as well as gene expression pattern

27. Pluripotent stem cells Germ cells Some adult stem cells
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Oocyte
Somatic cells
Nuclear
Transfer
Somatic cells
Fusion
Blastocyst
ES cells
Defined
factors
Culture
Pluripotent
stem cells
Germ cells
Some adult stem cells
Somatic cells

28. Therapeutic cloning
Produce patient-specific lines of embryonic stem cells
Artificial embryo created using same process as Dolly (SCNT)
Its cells are used as embryonic stem cells for transfer to injured tissue
Body readily accepts these cells with no immune rejection
May be obsolete with development of iPS cells

29. Pattern formation
All multicellular organisms seem to use positional information to determine the basic pattern of body compartments and overall body
Positional information then leads to intrinsic changes in gene activity
Cells ultimately adopt a fate appropriate for their location

30. Polarity – acquisition of axial differences in developing structure
Pattern formation can be considered the process of taking a radially symmetrical cell and imposing two perpendicular axes to define the basic body plan
Anterior–posterior (A/P, head-to-tail) axis
Dorsal–ventral (D/V, back-to-front) axis
Polarity – acquisition of axial differences in developing structure

31. Fruit fly Drosophila melanogaster
2 bodies during development
Larva – tubular eating machine
Adult – flying sex machine
Metamorphosis – passage from one body to next
Embryogenesis – process of going from fertilized egg to larva

32. Development begins before fertilization with construction of egg
Specialized nurse cells that help the egg grow move some of their own maternally encoded mRNAs into the maturing oocyte
Action of maternal, rather than zygotic, genes determines the initial course of Drosophila development

33. Syncytial blastoderm – 12 rounds of nuclear division without cytokinesis
Nuclei space themselves out
Membranes grow forming cellular blastoderm
Embryonic folding and primary tissue development soon follow
Within a day of fertilization, embryogenesis creates a segmented, tubular body

35. Anterior/Posterior axis in Drosophila
Based on opposing gradients of two different proteins produced by nurse cell mRNAs: Bicoid (anterior) and Nanos (posterior)
Morphogens – proteins whose concentration gradients can specify different cell fates
2 other maternal messages, hunchback and caudal, are evenly distributed across the egg
Bicoid inhibits translation of caudal mRNA
Nanos inhibits translation of hunchback mRNA

37. Pattern formation in Drosophila along the A/P axis
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Thorax
Head
Abdomen
Pattern formation in Drosophila along the A/P axis
Determination of structures is accomplished by the sequential activation of three classes of segmentation genes
Create body plan of three fused head segments, three thoracic segments, and eight abdominal segments

38. Within 3 hr after fertilization, a highly orchestrated cascade of segmentation gene activity transforms the broad gradients of the early embryo into a periodic, segmented structure with A/P and D/V polarity
Bicoid
Hunchback and other gap genes
Pair-rule genes
Segment polarity genes

39. 39 A T H Establishing the Polarity of the Embryo
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Establishing the Polarity of the Embryo
Setting the Stage for Segmentation
Fertilization of the egg
triggers the
production of Bicoid
protein from maternal
RN A in the egg. The
Bicoid protein
diffuses through the
egg, forming a
gradient. This
gradient determines
the polarity of the
embryo, with the head
and thorax developing
in the zone of high
concentration (green
fluorescent dye in
antibodies that bind
bicoid protein allows
visualization of the
gradient).
About 21/2 hours after
fertilization, Bicoid
protein turns on a
series of brief signals
from so-called gap
genes. The gap
proteins act to divide
the embryo into large
blocks. In this photo,
fluorescent dyes in
antibodies that bind to
the gap proteins
Krüppel (orange) and
Hunchback (green)
make the blocks
visible; the region of
overlap is yellow.
500 µm
500 µm
Laying Down the Fundamental Regions
Forming the Segments
About 0.5 hr later , the
gap genes switch on
the “pair-rule” genes,
which are each
expressed in seven
stripes. This is shown
for the pair-rule gene
hairy . Some pair-rule
genes are only
required for
even-numbered
segments while
others are only
required for odd
numbered segments.
The final stage of
segmentation occurs
when a “segment-
polarity” gene called
engrailed divides each
of the seven regions
into halves, producing
14 narrow
compartments.
Each compartment
corresponds to one
segment of the future
body . There are three
head segments
(H, bottom right), three
thoracic segments
(T, upper right), and
eight abdominal
segments (A, from top
right to bottom left). A T H 39
500 µm
500 µm
a: © Steve Paddock and Sean Carroll; b-d: © Jim Langeland, Steve Paddock and Sean Carroll

40. Drosophila mutants with particular segments that seem to have changed identity
Mutations in homeotic genes lead to the appearance of perfectly normal body parts in inappropriate places

41. Homeotic gene complexes of Drosophila
Bithorax complex
Several homeotic genes map together to 3rd chromosome
Control the development of body parts in the rear half of the thorax and all of the abdomen
Order of genes corresponds to order of segments
Antennapedia complex
Governs the anterior end of the fly
Order also corresponds to order of segments

42. a. b. 42 Drosophila HOM Chromosomes Mouse Hox Chromosomes
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Drosophila HOM Chromosomes
Mouse Hox Chromosomes
Drosophila HOM genes
Hox 1
Antennapedia complex
Bithorax complex
Hox 2
lab pb
Dfd
Scr
Antp
Ubx
abd-A
abd-B
Hox 3
Hox 4
Head
Thorax
Abdomen
Fruit fly
embryo
Mouse
embryo
Fruit fly
Mouse a. b. 42

43. Homeodomain
All of the homeotic gene complexes contained a conserved sequence of 180 nucleotides coding for a 60-amino-acid, DNA-binding domain
Homeobox DNA that encodes homeodomain
Hox gene
Homeobox-containing gene that specifies the identity of a body part
Function as transcription factors that bind DNA using their homeobox domain
Ultimate targets of Hox gene function must be genes that control cell behaviors associated with organ morphogenesis
Found in organisms as primitive as cnidarians

44. Evolutionary split between plant and animal cell lineages occurred about 1.6 BYA
Before the appearance of multicellular organisms with defined body plans
Multicellularity evolved independently in plants and animals
Genetic control of pattern formation in plants is fundamentally different from that of animals
Meristems add modules throughout lifetime

45. Plants have MADS-box genes
Homeotic gene family
Have homeobox-containing genes
Do not possess complexes of Hox genes similar to the regional identity ones in animals
Family of transcriptional regulators found in most eukaryotic organisms
Plants have many; animals only a few
In flowering plants, control transition from vegetative to flowering growth, root development, floral organ identity

46. Morphogenesis Generation of ordered form and structure
Product of changes in cell structure and cell behavior
Animals regulate
The number, timing, and orientation of cell divisions
Cell growth and expansion
Changes in cell shape
Cell migration (not used in plants)
Cell death

47. Orientation of the mitotic spindle determines the plane of cell division in eukaryotic cells
Plane determined by spindle placement
Great diversity of cleavage patterns in animal embryos is determined by differences in spindle placement
In animals, cell differentiation is often accompanied by profound changes in cell size and shape
Neurons, muscle cells

48. Apoptosis Programmed cell death a part of development
Human embryos begin with webbed fingers
Necrosis – cells that die due to injury
In C. elegans, due to 3 genes
Mechanism of apoptosis appears to have been highly conserved during the course of animal evolution
C. elegans genes similar to human genes

49. Cell migration important during many stages of animal development
Involves adhesion and loss of adhesion
Cell-to-cell interactions are often mediated through cadherins
Also involves cell-to-substrate interaction
Cell-to-substrate interactions often involve integrin-to-extracellular-matrix (ECM) interactions
Extracellular matrix controls extent or route of migration

50. Cadherins Large gene family with several subfamilies
All are transmembrane proteins that share a common motif
Cadherin domain
110-amino-acid domain in the extracellular portion of the protein that mediates Ca2+-dependent binding between like cadherins (homophilic binding)
Cells with the same cadherins adhere specifically to one another, while not adhering to other cells with different cadherins

51. Integrins
Attached to actin filaments of the cytoskeleton and protrude out from the cell surface in pairs, like two hands
“Hands” grasp a specific component of the matrix
Actions
Provides anchor
Can initiate changes in cell
Alter growth of cytoskeleton
Activate gene expression
Gastrulation depends on fibronectin–integrin interactions