1.2 Fertilization Flashcards
Calcium release from the ER, What does it do?
DAG and Ca2+ activate Na-H+ exhange pump
Increased Ca2+ leads to downstream
-cortical granules exocytosis
> slow block
>formation of hyaline
-activate NAD+ kinasse-->convert NAD+ to NADP+-->membrane biosynthesis
-Na in H+ out changes alkalinity
-changes that support embryo
>stimuklation of proetin synthesis, DNA replcation, cytoplastim movements
IP3
DAG
PIP3
H2O2
inositol 1,4,5 triphosphate
diacylglycerol
Phosphatidylinositol (3,4,5)-trisphosphate
hyrdrogen peroxide
OVERVIEW:
Sequence of events in sea urchin fertilization - 7
- Sperms swim towards egg: Chemotaxis
- Jelly coat stimulates acrosomal reaction.
- Sperm digest through jelly coat.
- Acrosomal process binds to vitelline envelope
- Fusion of egg and sperm membranes occurs;
depolarization of egg plasma membrane precludes further
sperm binding [fast block to polyspermy]. - Cortical reaction in eggs: fertilization envelope forms
more completely preventing sperm binding [slow block to
polyspermy]. - Fusion of egg and sperm pronuclei.
- Cleavage.
OVERVIEW:
The sequence of events in mammalian fertilization - 7
- Capacitated sperm (i.e. those that have had their surface
proteins/lipids cleaved or modified in the female reproductive
tract) wriggle and digest their way through follicle cells [or
cumulus oophorus), and bind to the zona pellucida (ZP). - Sperm undergo the acrosomal reaction.
- Sperm start to digest through ZP.
- Sperm membrane fuses with egg membrane [further sperm
binding precluded by a mild depolarization of egg membrane
i.e. “fast” block to polyspermy. - Cortical reaction occurs in egg-ZP is modified so it cannot
bind sperm; this is called the zona reaction [i.e., a “slow” block
to polyspermy]. - Fusion of egg and sperm pronuclei.
- Cleavage.
Sperm Capacitation
- remove cholesterol from cell membrane (changes fluidity of membrane)
- influx of HCO-3 + CA2+
- activate adenylate cyclase
- AC turns AMP–> cAMP
- activate PKA
- phosphoralate TK –>PTK
- phosphoralates sperm protions
- capacitation
- occurs in fem reproductive tract where sperm are active for 6 days
capacitated sperm can sense thermal gradient between
isthmus of oviduct and ampulla (about 2oC).
thermotaxis
sperm travel far. fluids in fem repro tract. fluid flux, sperm move against flow.
Rheotaxis
Zona Pellucida proteins in mice Vs. Humans
Mice: ZP1 ZP2 ZP3
Humans: ZP1 ZP2 ZP3 ZP4
if you block certain proteins in the egg or sperm, you block it and the interaction is hampered and the sperm will not bind to the egg
EXPERIMENT:
purified ZP and extracted protein then a sperm competition. What happened?
-purified ZP w egg and eperm mixutre
-ZP binds to the egg and prevents the sperm binding to it
-what interacts w egg cell surface? ZP1 ZP2 ZP3
-do again with solutions w sperate ZP proteins
-Assays show that ZP3 is most effective in
inhibiting sperm binding
-ZP3 is receptor to sperm on egg surface
-if spripped of carbohydrates ZP3 only some inhibition, therfore carbs are important
EXPERIMENT:
urified ZP and extracted protein then a sperm competition. What happenes in HUMANS?
- labeled acrosomal region green
- when rxn occurs the green disappers
- sperms had already done acrosomal rxns by the time they were first seen in cummulis
- took egg added ZP4 (human) to mouse oocyte and treated with human sperm
- ZP2 was the only one that interacts
- ZP2 binds to sperms in humans
- ZP2 binds onto mamalioan egg
ZP2 is cleaved by cortical granule protease ovostatin.
Mammalian Sperm Entry into Egg
- happens on the side
- The junction between the inner acrosomal membrane and sperm cell membrane is called Equatorial region.
- Through the equatorial region the fusion with egg cell membrane is initiated.
the sperm is in the egg, now what?
complex cascade of events, largely centered
around Ca++ release from ER storage:
• activation of protein synthesis–from stored
mRNA–”maternal” messenger RNA
• activation of DNA synthesis
• increase in oxygen consumption
• migration of sperm and egg nuclei toward each other
egg is now said to be “activated”
• can be imitated (up to a point) with Ca++
ionophores, or even a simple mechanical
stimulus–parthenogenesis (fake activation)
Fusion of genetic material
Sperm’s nucleus and centriole separate from the mitochondria and
flagellum.
• The sperm nucleus undergoes dramatic decondensation to form
pronucleus.
• Nuclear envelope vesiculates into small packets, and the sperm’s proteins
associated with condensed chromosomes are exchanged by egg’s to
decondensate chromosomes.
• 180° rotation of male pronucleus results in the localization of centriole
between male and female pronucleus.
• Centrioles act as microtubule organizing center integrating both egg
microtubles with sperms.
• Male and female pronuclei migrate towards each other and their fusion
results in the formation of diploid zygote.
• Phosphorylation of lamin and histones by PKA after the binding to
glycoprotein in the egg jelly.
Formation of the first mitotic spindle (in
mammals
• pronuclei migrate toward each other
• male pronucleus decondenses (under the
influence of egg proteins); nuclear envelope
vesiculates; sperm centrioles mature
• female pronuclear envelope also vesiculates
• sperm centrioles replicate
• first spindle is formed
• mitosis occurs: 1st cleavage or division (in
mammals, at about 12 hours, in sea urchin,
about 1.5 hours)
Genomic imprinting
In mammals there are ~100 genes where the active
allele is either maternally derived or paternally
derived, but not both.
• This phenomenon seems to be due to the fact that
during gametogenesis, some genes are methylatedthus
bear an imprint on their DNA.
• The pattern of methylation can be different in sperm
and eggs: e.g. an allele is methylated in eggs, not
sperm, or vice versa.
• At fertilization, the zygote contains one allele which is
active and one which is not.
• This pattern is erased and “reset” in the next
generation during gametogenesis
