Unit 4 Fertilization What Does Fertilization Accomplish Sexual reproduction allows meiotic recombination genetic variation Restores diploid genome Sperm brings in a paternal centrosome organizes microtubules for pronuclear migration and first mitosis Species specific sperm egg union Leads to blocks to polyspermy Egg activation includes completion of meiosis in many species Fertilization Questions to Answer What activates stimulates sperm What helps to confer species specific sperm egg binding What confers the fast and slow permanent blocks to polyspermy What cause the sperm to undergo the acrosome reaction Sea urchin fertilization Gilbert 8e Fig 7 14a cf 10e Fig 4 14a Sea Urchin Egg Structure Egg jelly produces molecules that attract sperm Sperm must digest it Gilbert 10e Fig 4 4 The vitelline envelope is attached to the egg plasma membrane Sea Urchin Sperm Undergo Chemotaxis Toward RESACT Chemotaxis moving toward higher concentrations of a molecules the chemoattractant RESACT is a peptide released from egg jelly Kirkman Brown et al 2003 Nat Cell Bio 5 93 96 Sea Urchin Sperm Undergo Chemotaxis Toward RESACT 0 sec 20 sec 40 sec 90 sec Gilbert 10e Fig 4 9 Sea urchin fertilization Egg jelly triggers the acrosome reaction Gilbert 10e Fig 4 8 Sea urchin acrosome reaction Gilbert 10e Fig 4 12 Sea urchin acrosome reaction Gilbert 10e Fig 4 12 Sea urchin sperm contain bindin From Gilbert 3e Fig 22 17 Bindin is on the surface of the acrosomal process Kalthoff 1e Fig 4 7 Question Calcium is sufficient to trigger the acrosome reaction What would we need to test this Calcium ionophores such as A23187 and ionomycin are drugs that trigger release of calcium in the absence of normal signals Sea urchin fertilization timeline The fast block occurs within milliseconds to a few seconds The slow block occurs on the timescale of minutes Fast Sperm into egg block fert membr Sperm begins Slow formed Nuclei block attachfuse begins ment 1 10 100 1000 1st cleavage Log sec DNA synth pH i incr starts respir incr Incr protein synth Ca 2 i incr Fast block to polyspermy involves membrane depolarization Gilbert 10e Fig 4 17 Slow block to polyspermy sea urchins Cortical granules release proteases that release the envelope Gilbert 10e Fig 4 19 Cortical granules Gilbert 3e Fig 2 6 Slow block to polyspermy sea urchins Cortical granules also release enzymes that result in hardening of the envelope and proteoglycans that swell aiding lifting of the envelope Gilbert 10e Fig 4 19 Sea urchin bindin is species specific Gilbert 910e Fig 4 13 Sperm egg fusion the fertilization cone draws sperm in fertilization cone Courtesy Jerry Schatten Testis Organization Sperm maturation occurs in the epididymis In order to fertilize efficiently capacitation must occur in the female reproductive tract adapted from Gilbert 10e Fig 17 28 Mammalian sperm undergo capacitation in the oviduct Gilbert 10e Fig 4 29 Review Mammalian Oocyte Structure Purves et al 6e Fig 43 2 Mouse fertilization The cumulus layer partially activates sperm the zona triggers the full acrosome reaction Gilbert 10e Fig 4 8 Mouse acrosome reaction Gilbert 10e Fig 4 30 Mammalian sperm do not make an acrosomal process The acrosome reaction probably begins during contact with the cumulus layer no acrosomal vesicle green intact acrosomal vesicle Gilbert 10e Fig 4 31 The zona pellucida is a key in mammalian fertilization P Wassarman The zona pellucida is fibrous Paul Wassarman The zona pellucida contains three main proteins ZP3 is glycosylated sugars groups are attached to specific amino acids Gilbert 7e Fig 7 17a A competition assay can be used to identify key zona proteins sperm added oocytes added zona pellucida Kalthoff 2e Fig 4 11 A competition assay can be used to identify key zona proteins sperm added oocytes added Kalthoff 2e Fig 4 11 zona protein blocks sperm binding Mammals ZP3 Competition Conclusion glycosylated ZP3 is important for sperm zona binding Gilbert 9e Fig 4 32a Zona remodeling and the slow block The zona is enzymatically remodeled after fertilization This contributes to the slow block to polyspermy in mammals Gilbert 9e Fig 4 31 ZP2 remodeling may underlie the slow block Lots of sperm still attaching sperm DNA sperm tails Mutant ZP2 that can t be cleaved does not allow slow block Gilbert 10e Fig 4 33 Mammals Sperm fusion Gilbert 10e Fig 4 32 What about sperm oocyte binding After decades of study the answer is still unclear Probably involves multiple receptor ligand interactions See your text for discussion