Bio 127 - Section III Late Development

Bio 127 - Section IIILate Development

Germ Line DevelopmentGilbert 9e – Chapter 16

Section 4 Encompasses :

• Development of the Tetrapod Limb

• Sex Determination

• The Saga of the Germ Line

• Post-Embryonic Development

Student Learning Objectives

1. You should understand that sexual reproduction requiring the fusion of gametes from male and female gonads occurs in specific organisms.

2. You should understand that the primordial germ cells that give rise to gametes arise outside of the gonads and must migrate to them.

3. You should understand that in most organisms the primordial germ cells are specified conditionally, while in some they are specified autonomously by cytoplasmic determinants in the egg.

4. You should understand that migration of the germ cells from their site of origin to the gonads is an essential part of reproductive success .

• In all plants and some animals, somatic cells can readily form new organisms– Cnidarians, flatworms, tunicates

• In many animals, there is an early division between somatic and germ cells– Insects, roundworms, vertebrates

• Two step process;

– Primordial germ cells (PGCs) are determined in a specific location in the embryo

– PGCs migrate to the gonad and become the progenitor population for eggs and sperm

Two Methods of Germ Cell Determination

• Autonomous Specification– Egg cytoplasmic determinants– Called ‘Germ Plasm’– Nematodes, flies, frogs

• Conditional Specification– Signals from surrounding cells– Majority of sexually reproducing organisms– Including mammals

The nematode Caenorhabditis elegans

Remember cleavage and gastrulation:Asymmetrical divisions produce astem cell (P-lineage), “founder” cell.

Stem cell divisions are meridional

Founder cell divisions are equatorial

Gastrulation in C. elegans

P-granules hold cytoplasmic determinants in C. elegans

Blue is DNA marker, Green is P-granule marker

P4

PIE-1: Blocks all transcription, thus all differentiationGerm plasm also has blocks to translation, stem cell factors, controls for asymmetric divisions and meiosis inducing agents.

Blue stain marks transcriptional activity

P4

Synctitial cleavage in Drosophila is followed by cellularization

Pole plasm forms during cellularization

anterior posterior

- mitochondria- fibrils- polar granules no transcription no translation germline stabilization

Localization of germ cell-less (gcl) gene products

Human males with mutant homolog are often sterile

Germ plasm at the vegetal pole of frog embryos

Markerfor froghomologof fly/wormtranslationblocker,Nanos

• The frog cells that take up these granules will become PGCs and migrate to the gonads as the kidney forms– Again, no transcription or translation– Therefore, no differentiation

Conditional Specification of mammalian PGCs

• Posterior of epiblast at the junction of the primitive streak and extraembryonic ectoderm– Cells are no different from other epiblast– Specified in gastrulation before 3 layers form– Wnts from endoderm make them competent– BMPs from extraembryo ectoderm finish it

Picture the blastocyst full of yolk.....

Poor old Henson discovered this node as well but didn’t get the naming rights

Conditional Specification of mammalian PGCs

• Same deal as the others:– Repress differentiation by repressing gene

expression

• Specified outside embryo forming cells– Once expression is shut down they can go

back into embryo and not respond to signals

Germ Cell Migration

• Drosophila

• Zebrafish

• Frogs

• Mice

• Birds and Reptiles

Germ Cell Migration: Drosophila

As the endoderminvaginates, the ectoderm andmesoderm extendand converge towrap around thedorsal side to formthe “germ band”


anterior posterior

- mitochondria- fibrils- polar granules no transcription no translation germline stabilization


Germ cellspassively rideendoderm


Endoderm expressesrepellent molecules

PGCs and gonadprogenitors in 2 migration streams

Germ bandis retracting


Combination of chemoattractionand repulsion drive them to gonad

E-cadherin METforms epitheliumaround PGCs


• Both mesoderm and PGCs divide through the larval stage, differentiate at metamorphosis

• At larval-pupal transition anterior PGCs in gonad become germ line stem cells

• In ovaries, the cells attach top stromal cap In testes, the cells attach to hub cells

Remember: Zebrafish development occurs very rapidly

24-hours from 1 cell to vertebrate embryo!

Germ Cell Migration: Zebrafish

specification: germ plasm determination: PGCs by 32-cells

four clustersjoin into two

migration of bilateral clusters into developing gonad follows signal Sdf-1 using receptor CXCR4

Remember: Germ plasm at vegetal pole in frogs

Markerfor froghomologof fly/wormtranslationblocker,Nanos

Germ Cell Migration: Frogs

During cleavage the germ plasm rises up until it ends up in the endoderm at top and back near lip


The endoderm belowmesdoderm are PGCs


Migration anterior to gonads atendoderm-mesoderm boundary

~30 PGCs reach gonadsby fibronectin and Sdf-1

• Remember Sdf-1

• Soluble signal whose receptor is CXCR4

• Common signal for vertebrate germ cells

• Also used by humans to call HSC to bone marrow, guide lymphocytes, MSC?

Germ Cell Migration: Mice

PGCs formed in extraembryonic epiblast

10-100 cells@ Day 6.5

in mice


Once formed, they migrate directly into the hindgut endodermand migrate anteriorly through Day 9 dividing the entire time

They leavethe gut bythe dorsalmesentaryand enterthe genitalridges by Day 12 as2500-5000PGCs.


• The travelling stem cell niche

– Support cells travel with PGCs to maintain the undifferentiated stem cell phenotype

– They secrete stem cell factor (SCF)

– The cells follow fibronectin trail

– Sdf-1 also required

Germ Cell Migration: Birds and Reptiles

Germ line cells determined in thearea pellucida, migrate to hypoblast

Migrate to gonads via blood streamwhen extraembryonic vessels form

Germ Cell Migration: Birds and Reptiles

Sdf-1 from intermediate mesodermdraws them out of vessels and throughthe mesodermal tissues to the gonad

Bio 127 - Section IIILate Development

Post-Embryonic DevelopmentGilbert 9e – Chapter 15

Section 4 Encompasses :

• Development of the Tetrapod Limb

• Sex Determination

• The Saga of the Germ Line

• Post-Embryonic Development

Student Learning Objectives

1. You should understand that development never stops during the life of the organism and that three major processes occur in the post-embryonic animal: metamorphosis, regeneration and aging.

2. You should understand the Direct Development involves young organisms with the same body plan as the adult; whereas Indirect Development involves major changes to form the adult body plan.

3. Indirect Development, or metamorphosis, is hormonal reactivation of

4. You should understand that regeneration is the reactivation of developmental process to restore missing tissues.

5. You should understand that aging and physiological senescence are an interplay of genetic and environmental influences.

Metamorphosis

• Development of a larval stage and an adult stage specialized for different functions– Larvae often specialized for growth, dispersal, etc.– Adults usually specialized for reproduction– Example Cecropia moths:

• Larvae are wingless eating machines• Adults have one day to mate – don’t even have mouth parts!

• Two major types of larvae– Primary: little to no similarity to adult (sea urchins)– Secondary: add and subtract parts from similar form

• (insects, amphibians)

Metamorphosis: Sea Urchins

PluteusLarvae

Metamorphosis: Sea Urchins

PrimaryLarvae:No traceof adultmorphology

Metamorphosis: Amphibians

• Hormone(s): T3 and T4

• Four Major Morphological Processes– Growth of new structures– Cell death in existing structures– Remodeling of existing structures– Biochemical respecification

• Shift in the genes expressed and the physiological functions they control


New neuronsdifferentiateand form newipsilateral tracts

Tadpole eyes areon the sides of the head, frog eyes areon the front and top

Binocular Vision


• Cell death in existing structures

– Apoptosis• T3 induces apoptosis in tail and gills• Apoptosis occurs in gut epithelium due to ECM loss

– Phagocytosis• Macrophages finish off the cells of the tail• Also destroy larval RBCs to make fresh ones with

the adult hemoglobin protein


Remodeling: - Eyes - Skull - Skeleton - Gut - Sensory


Biochemical Respecification

NH3 = ammonia, amino groupNH4

+ = ammonium ion

T3 causes a shift in transcription factor expression that upregulates these genes.


2NH3 + CO2

+ H2O

(urea)

Actions of thyroxine (T4) and tri-iodothyronine (T3)

Thyroid receptors arefound in the nucleuswhere they are boundto inactive promoters.

When thyroid hormoneenters such a cell, it willbind to receptor and thecombination is an activetranscription factor forthat specific gene.

Cells that make highlevels of deiodinase IIare more responsive tothyroid stimulation.

Cells that make highlevels of deiodinase IIIare less responsive

Actions of thyroxine (T4) and tri-iodothyronine (T3)

• What genes you have your thyroid receptors on is very important to your function– Limb muscle cells grow in response to thyroid hormones– Tail muscle apoptoses in response to thyroid hormones

• How much deiodinase II you make is very important– Limb buds make a lot and respond to early low levels of T4

– Tails make very little and wait for later very high levels of T4

– This is good!

• How a tissue is organized before T4 is very important– Thyroid hormones make skin apoptose– Head and body have basal stem cells, tail does not

• Your tail degenerates during week 4 of gestation in much the same fashion as the frogs!


• Some amphibian species have evolved alternatives to metamorphosis: Heterochrony

– Neoteny: Normal gonadal maturity, retention of juvenile form

– Progenesis: Accelerated gonadal maturity, retention of juvenile form

– Direct Development: No larval form


Neoteny in the Mexican axolotl (salamander)

Normal adult with juvenile form.

Adult form not seen in nature,resulting from T4 in the pond.


Direct development in a common Puerto Rican frog

Two views of the developing limb buds within the egg

Frogs, not tadpoles,hatch from the eggs.

Very large, nutrient-rich eggs allowskipping larval food gathering stage.

Metamorphosis: Insects

Adult = imago


• Larva are eating machines to provide energy for non-feeding pupal development

• They have both doomed larval cells and rudiments of imaginal cells for adult

– The larval cells will apoptose in the pupa

– Imaginal disc cells will form wings, legs, antennae, eye, head, thorax and genitalia

– Histioblasts will form adult abdomen

– Imaginal cell clusters in each organ will form adult organ as the larval organ degenerates


Locations and developmental fates of imaginal discs and imaginal tissues in the third instar larva of Drosophila


Leg Imaginal Disc

beforepupa

duringpupa

Epidermis cellsform a hollowtube that coils

Telescopesout in pupa



A view into the minds of fly-guys....

The cells at center of the discsecrete Wingless (Wnt) andDecapentaplegic (TGF-B)

This causes differentexpression levels oftranscription factorsDachsund (green) andDistal-less (red).


• Like amphibians, control is hormonal in insects

– Presence of juvenile-hormone makes a larval molt

– Shift to steroid 20-hydroxyecdysone gives pupal molt

– Differential timing in the development of pupal structures is due to 20E receptor expression timing

Regeneration

• Restoration of missing tissues

• Post-embryonic reactivation of development

• Occurs in some form in all species

– Stem cell-mediated regeneration

– Epimorphosis: adult cell de- and re-differentiation

– Morphallaxis: adult cell repatterning

– Compensatory regeneration: adult cell division

Regeneration: Epimorphosis

flatworms,salamanders

The survivingcells lose theirspecificationand form anundifferentiatedtissue bulge

Regeneration : Epimorphosis

Vertebrate limb development from apical ectodermal cap (ridge)

Regeneration: Morphallaxis

The hydra is constantly regenerating cells that are sloughed off.

Only the head and foot aren’t moving


• Any cells along the length can become head, body or foot

• If you cut a hydra up all pieces will form all structures

• Each piece has a dual gradient already established and both ends are specified


The bud always forms roughly the same distance anterior-posterior due to the combined gradient

Regeneration: Compensatory Regeneration

• No dedifferentiation occurs in liver regeneration

• All five cell types produce more of themselves• hepatocytes• duct cells• fat-storing (Ito) cells• endothelial cells• Kupffer macrophages

• Progenitor cell back-up plan: Oval cells

Aging

• Time-related deterioration of physiological functions necessary for survival and fertilization

– Life span vs. senescence

• Combination of:

– Mutations

– Environmental factors

– Random epigenetic changes

Aging

We’ve learnednot to die youngas often more than we’ve extended life

Aging

Mutation repair deficiency

Hutchinson-Gilford progeria

Aging

Low caloric intakeis associated withlong-life in all species.

Figure 15.36 Differential DNA methylation patterns in aging twins (Part 1)

Epigeneticalterations

Figure 15.37 Methylation of the estrogen receptor gene occurs as a function of normal aging

Can effectreproductivecapacitydirectly

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Bio 127 - Section III Late Development