Developmental biology - understanding the mechanisms that control embryo development
Understanding which genes are turned on when which are turned off & how its regulated
Differentiation
Gene expression - genes that are actively transcribing mRNA
Genetic equivalence - genetic material in almost all cells is the same - exception is immune cells that have more variation in their DNA for generating antibodies
Evidence = Sir John Burden 1960s - nucleus of a frog's oocyte replaced with nucleus from gut cell - tadpoles can still be produced
Different distinct cell phenotypes occur because only a proportion of the genes are expressed (eg. neurons, pancreatic)
Somatic cells:
Cells of the body
Limited life
Germ line cells:
Cells for reproduction
Immortal
Female - oocytes
Male - sperm
GERM LINE
Oocyte and sperm cells generated from primordial germ cells
Oogenesis
Meiosis
Haploid DNA content
Large gamete
Spermatogenesis
Meiosis
Small motile gamete
Haploid DNA
Mammalian development
Most development knowledge comes from studying mice - easy to manipulate
Monotremes - lay eggs
Marsupials - shorter gestation, immature young
Placentals - placentas, more mature when born than marsupials
Mouse embryo development:
Human embryos don't undergo the elongation - they remain as a flat disc
Why don't all the epiblast cells become primordial germ cells?
Cell signalling
Bone morphogenetic protein 4 (BMP4) - signalling molecules secreted from extraembryonic tissue
BMP4 acts on cells in the epiblast that its most closely in contact with - it triggers the cells to become PGCs
High levels of BMP4 are needed for th
BMP4 receptors:
Transmembrane proteins
Extracellular part is where the BPM4 binds
Binding causes phosphorylation in cytoplasmic part - leads to cascade - new genes in the nucleus are turned on
How do we know this?
Cells from the top of the epiblast were transferred and transplanted to the region closest to the extraembryonic tissues
These cells are exposed to high enough levels of BMP4 to become PGCs
Shows that fate of the epiblast cells depends on their position in the embryo
Differentiation of PGCs is controlled by the environment
Migration of PGCs:
Mechanism of their migration not fully understood
They’re generated in the epiblast on day 6-7
Day 8 - migration into the gut
Day 11 - enter the genital ridges (tissue that gives rise to ovaries and testes)
6PGCs at the start - around 5000 when they enter the genital ridges
Differentiation into sperm or oocytes?
PGCs in female genital ridge are committed to an oocyte fate and vice versa
XY PGCs in a foetal ovary will develop as oocytes
XX PGCs in foetal testis will develop as sperm
Anne McLaren
Worked on PGC differentiation
- suggested that retinoic acid is important (metabolite of vitamin A)
IVF - showed the possibility to fertilise mouse oocytes in vitro to generate viable embryos
Retinoic acid signalling:
Retinoic acid:
Metabolite of vitamin A
Lipophilic - easily passes over the cell membrane
Important in stages of embryonic development by regulating gene expression
Potent
Present in female and male foetal gonads
Needed for PGCs to become oocytes
Male gonad cells express enzyme cytochrome P450 which degrades RA - inhibits oocyte differentiation
If XY PGCs are removed from the male gonad and placed in female gonad:
RA levels in environment increase, RA induces PGCs into oocytes
If XX PGCs are placed in male gonad - presence of cytochrome P450
RA degraded - PGCs differentiate into sperm
→ sperm
→ oocyte
Absence of retinoic acid - staining shows retinoic acid binding proteins on the outer area of the cell
Presence of retinoic acid - rapid translocation of the protein into the nucleus
SPERMATOGENESIS
Production of mature sperm cells from PGCs
Continuous and prolific
Occurs in seminiferous tubules of the testes
In male gonad, PGCs give rise to stem cells called spermatogonial sperm cells (SSCs)
SSCs generate more SSCs and differentiate to form primary spermatocytes
Primary spermatocytes undergo meiosis to become secondary spermatocytes
Secondary spermatocytes give rise to spermatids
SSC proliferation and differentiation needs to be tightly controlled
Too MUCH proliferation could lead to formation of a tumour
Too LITTLE proliferation and too MUCH differentiation could deplete the SSC population - leading to male infertility
Function of sperm components:
Acrosome - derived from golgi, contains enzymes that digest proteins and sugars; these enzymes are required to lyse the outer coverings of egg
Nucleus - contains haploid number of chromosomes
Midpiece - contains mitochondria that produce the energy (ATP) required for motility
Flagellum - required for propulsion, motor portion of the flagellum is the axoneme
OOGENESIS
Development of mature oocytes from PGCs
Occurs in ovaries
Begins in the embryo with the differentiation with the PGCs - to stem cells called oogonia
Oogonia multiply by mitosis and begin meiosis - stops at prophase 1 - primary oocytes
Primary oocytes remain arrested in prophase 1 until puberty
Each month follicle stimulating hormones trigger division of some of the primary oocytes
First mitotic division produces uneven sized cells - one is secondary oocyte, one is polar body
Secondary oocyte begins second mitotic division but is arrested in metaphase 2
If fertilised by sperm - complete second division - gives rise to 2nd polar body
Once meiosis 2 is complete - nucleus of ovum fuses with sperm nucleus - forms a zygote
Structure of oocyte:
Nucleus - nucleus of mature oocyte is arrested in 2nd metaphase
Zona pellucida - thich extracellular matrix that binds sperm
Cumulus - layer of ovarian follicular cells surrounding oocyte, layer adjacent to zona called corona radiata
Cytoplasm - contains proteins, ribosomes, tRNA, mRNA
FERTILISATION
Attraction and activation of sperm by contents of female reproductive tract
Different regions of tract secrete molecules that attract and affect sperm motility
In some mammals, sperm becomes hyperactivated in the oviduct
Ovarian follicle may secrete chemo-attractants that attract sperm towards oocyte
Binding of sperm to zona pellucida
Binding is species specific eg. mouse sperm only bind to oocyte that contain ZP3 glycoprotein on their surface
Release of enzyme from acrosome to lyse hole in zona
Eg. in mice - acrosomal reaction triggered by cross linking of proteins on sperm surface to ZP3
Enzymes released to make hole in zona so sperm can reach plasma membrane of the ovum
Passage of sperm through zona
Plasma membranes of sperm and ovum fuse
Sperm nucleus enters
Fusion of sperm and oocyte pronuclei
Sperm entry - female pronucleus stimulated to complete its 2nd meiotic division
Chromatin of male pronucleus uncoils
Each pronucleus migrates towards the other
Two nuclear envelopes break down
Chromosomes orientate themselves on the mitotic spindle - creating a zygote
→ female and male pronuclei are not equivalent
Some genes are imprinted and only expressed from either maternal or paternal chromosome
Hydatiform mole - only has male chromosomes - mass of placenta like cells - embryo doesn't develop - can give rise to tumours
Parthenogenetic embryos - only has female chromosomes - sometimes has organs - chaotic development and embryo becomes grossly disorganised
Prevention of polyspermy:
After fusion - cortical reaction occurs
Egg releases enzymes that harden zona pellucida so no more sperm can penetrate
Mice - enzymes modify ZP3 so oocyte can't bind to sperm