Advanced Glycobiology May 2016
Glossary of Terms for Fertilization. Implantation, Placentation
Accessory Gland (male reproductive): Prostate, seminal vesicle and Cowper’s
(bulbourethral) gland, that contribute to seminal fluid during ejaculation.
Amniotic Fluid: Fluid bathing the developing embryo.
Acrosome: Sperm organelle derived from Golgi.
Alpha agglutinin: Mating type protein in yeast.
Anisogamy: Gametes differ in size.
ART: Assisted Reproduction Technology.
Atresia: Degeneration of ovarian follicles resulting in depletion of follicles with
eggs in females.
Beta-Lactoglobulin: Major whey protein in cow and sheep’s milk.
Beta Defensin 126: Sialylated epiddymal defensing with mucin-like domain.
Bindin: Sperm protein in sea urchin sperm.
CA-125: (MUC 16) Mucin found on immune cells and secreted by ovarian cells,
both normal and cancerous.
Capacitation: Changes in sperm triggered by exposure to female tract factors
resulting in membrane reorganization motility and signal transduction.
Chauvinist gene: Gene only expressed in the male germ line e.g. protamines,
DEF126, PH20.
Corpus luteum: (yellow body) Temporary endocrine structure formed by
ovulated follicle in female ovary.
Cortical Reaction: Degranulation reaction releasing glycoproteins and enzymes
that block polyspermy upon fertilization.
Cumulus Oophorus: Mass of cumulus cells and secretions associated with the
ovulated egg. The cells secrete hormones (progesterone) and have been
suggested to be involved in nutrition of sperm, selection of sperm with normal
morphology, induction of acrosome reaction and support of oocyte maturation.
Cytotrophoblast: Placental cells attached to the placenta, forming the disk
shaped placenta and the villi.
Decidua: Lining of the endometrium which is shed each month.
Endometrium: Inner mucus membrane of mammalian of uterus.
Epididymis: Organ next to testes that is involved in sperm maturation.
Fallopian Tubes (Salpynx/oviduct): Upper portion of the female reproductive
tract.
Feto-Embryonic Protection System: Concept of molecular networks that shield
the fetus and embryo from maternal immune rejection.
Follicle (Ovarian): Structure holding female germ cells.
Follicle stimulating hormone (FSH): FSH is synthesized and secreted by
the gonadotropic cells of the anterior pituitary gland,[1] and regulates the
development, growth, pubertal maturation, and reproductive processes of the
body.
Fucoidan: Sulphated fucose polymer.
FUT5: Fucosyl transferase on sperm membrane that acts only as lectin.
Germ line/ Soma: Cells that generate gametes and cells that form the
perishable body.
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Advanced Glycobiology May 2016
Glycodelin (Placental Protein 14, PP14/ Progestagen associated
endometrial protein, PAEP): 180 AA lipocalin protein heavily glycosylated
17.5% by weight) with hoimology to β lactoglobulin.
Glycodelin A: Isoform with anti-fertilization properties secreted by female
reproductive tract. Immune suppressive, pro-apoptotic for immune cells and
involved in implantation.
Glycodelin S: Non-sialylated. Fucose-rich glycoform, found in seminal plasma.
Glycodelin C: Deglycosylated version of Glycodelin F (taken up by cumulus
cells and partially deglycosylted), stimulate sperm-ZP binding
Glycodelin F: Follicular fluid Gdn, binds spermatozoa in head region and
inhiibits acrosome reaction.
Granulosa Cells: Cells of the ovarian follicle that give rise to cumulus oophorus,
produce sex steroids and growth factors.
Hemizona Assay: Use of bisected matching halves of an ovum to test ZP
binding by capacitated sperm in vitro. One of the halves can be exposed to test
substance while the other provides the control.
Hyperactivation: Characteristic change in motility of sperm after capacitation.
ICSI: Intra-cytoplasmic sperm injection, allows fewrtilization of eggs by immotile
sperm, used in humans since 1992.
Isogamy: Gametes are of same size, ancestral state still in existence in some
algae and yeast.
IVF: In vitro fertilization, involving the overlay of an ovum with millions of sperm.
Interstitial trophoblast: “Free floating” invasive throphoblast (fetal placental)
cells that invade the decidua and interact with NK cells of the mother.
Luteinizing hormone (LH): Hormone produced by gonadotropic cells in the
anterior pituitary that triggers ovulation in females, stimulates testosterone
production by Leydig cells in males.
Myometrium: Muscular layer of the uterus.
Ovastacin: A metalloproteinase similar to hatching enzymes from arthropods
released by cortical granules, cleaves ZP2.
Perivitelline Space: Space between egg membrane and zone pellucida.
PH20/SPAM1: Sperm Adhesion molecule 1, HYAL1, hyauronidase 1.
Polycystic Ovary Syndrome (PCOS): Condition associated with increased
secretion of luteinizing hormone, androgen and insulin.
SialylLewisX: Tetra saccharide with α2,3 linked Sia and α1,3 linked fucose.
Sperm Chromatin Structural Assay (SCSA): Assay for quantifying DNA breaks
in sperm.
Syncytiotrophoblast: Outermost layer of placenta (trophoblast) consisting of
multinucleated cells (syncytia).
Trophoblast: Outer layer of blastocyst which gives rise to placenta.
Utero-Tubal Junction: Transition between uterus and lower fallopian tubes,
Vestment: Extracellular matrix and cumulus cells surrounding the ovulated
ovum.
VERL: Mollusk egg coat receptor for sperm lysine.
Window of Implantation: Eight days after luteinizing hormone surge (ovulation)
Zona pellucida: (“transparent zone”) Extra-cellular matrix of the mature egg.
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Advanced Glycobiology
Lecture 14
Fertilization, Implantation, Placentation
May 12, 2016 Pascal Gagneux
Fig. 1. (A) Life cycle of C. crescentus.
Caulobacter crescentus
Ackermann & Stearns Science (2003)
Advantages of Sex
Reproduction
Asexual
Parthenogenesis
Division, Reproduction, Aging?
Sexual
•  Purging of deleterious mutations.!
•  Maintenance of genetic variation.!
•  Evolutionary Evasion of pathogens and
Aphid fundatrix giving birth to already gravid offspring
parasites (Red Queen).!
Female Whiptail lizards laying unfertilized eggs
Most vertebrates
Disadvantages / Cost of Sex
Micro and Macro Gametes
•  Need for males and female!
•  Loss of co-adapted allele complexes!
•  Why are same size gametes so rare?!
•  Conflict between sexes!
•  Genomic conflicts, e.g. meiotic drive.!
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Requirement for Gamete Recognition Rapid Evolution of Recognition •  Costs of Outbreeding and Inbreeding!
The existence of species rests on a metastable equilibrium between inbreeding and outbreeding.!
Swanson & Vacquier. Nat. Rev. Genetics (2002)
Joly, E. Biology Direct (2011)
Asexual, Parasexual, Sexual External vs Internal Fertilization
•  How to find each other?!
•  How to survive in another individual’s body?!
Ene, J.V. and Bennett, R.J. Nat. Rev. Microbiol. (2014)
External vs Internal Gestation
Implantation
•  Laying eggs and incubation.!
•  Laying developing eggs (brood parasites, Honey
Guide)!
•  Embryo -Endometrial recognition (opportunity for
quality control).!
•  Ovo-vivipary (Tsetse Fly)!
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The Molecular Basis of Sex
Placentation
•  Immunological Paradox of Mammalian Pregnancy!
Human
ZP
Abalone
Yeast
VERL alpha aggluinin
Abalone
VERL with N-Glycosylation sites
Peter Medawar!
Swanson et al. Mol. Biol Evol. (2011)
Immunological Paradox of Mammalian Pregnancy!
Victim of parental glycan mismatch!
1956, Obstetrics and Gynecology
Immune paradox of insemination!!
• 
• 
Male Gametes!
Male germ cell must navigate to the egg!
- survive and evade female immune response!
- retain functional potential of acrosome!
- retain integrity of haploid genome!
Female must protect against incompatible sperm!
!
Species
Population
Individual
Gamete
• 
Correct species!
• 
Not a pathogen!
• 
High fitness mate!
• 
• 
Correct diploid
alleles!
Compatible
genome!
• 
Acrosome-intact!
• 
Genetically intact!
Darszon et al. Physiol. Rev. (2010)
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Lectins on Sperm
The Mammalian Sperm Glycome!
GPI anchored
glycoprotein
loosely associated glycoprotein
(e.g. glycodelin S)
gangliosides
polysialic acid
membrane glycoprotein
heparan sulfate
GPI anchored
glycoprotein
beta defensin
gangliosides
Nicolson & Yanagimachi Science (1972)!
beta defensin
Tecle & Gagneux 2015 Mol. Repr. Develop.
Synthesis of the Mammalian Sperm Glycome!
Spermatogenesis!
Tecle & Gagneux 2015 Mol. Repr. Develop.
Sperm Glycocalyx
Tecle & Gagneux 2015 Mol. Repr. Develop.
Sperm Glycocalyx
SPAG11!
CD52!
Schröter, S, et al. Human Repr. Update . (1999)
Schröter, S, et al. Human Repr. Update . (1999)
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The Epididymis, an extra-cellular golgi?!
Acquisition of a special, highly sialylated
glycocalyx during epididymal transit:!
CMP-Neu5Ac
Sugar nucleotide: CMP-Neu5Ac
Glycosyl transferases: ST6Gal I
0.16
DEFB126
highly sialylated glycoprotein
CD-52, PH20,CD55, CD59, Prion GP
sialylated GPI anchored gycopeptide
Gangliosides
(mono-. di- & trisialylated)
0.14
0.12
Picmol/
100ug
protein
0.1
0.08
0.06
0.04
0.02
0
Testis
Head1
Head2 Corpus
20 glycosylation sites/ DEFB126, heavily
sialylated
Cauda
Ma et al. in press
Gain of Sialic Acids
“SAMP”
Bellannée et al. 2013 Cell Tissue Res. Ma et al. in press
Tollner, T. L. et al. 2012. Nat. Rev. Urol.
Ma et al. in prep.
Self Associated
Molecular Pattern
DEF126 deletion mutation!
DEFB126!
Lectin binding of human sperm
associates with DEFB126 mutation
and serves as a potential biomarker
for subfertility, yet frequency is
around 45%!
Theodore L. Tollner, Charles L. Bevins & Gary N. Cherr
Nature Reviews Urology 9, 365-375 (July 2012)
Immunological Paradox of Mammalian Fertilization!
Xin et al. Scientific Reports (2016)
Female Reproductive Immunity!
Leukocytic Reaction
1
Length of
oviduct
~15cm
Opportunities for Sperm competition and
Cryptic Female Choice
Uterine cavity:
leukocytes antibodies
complement
Oviduct
CAPACITATION
100s
Cervical crypts
~15cm
60 µm
30 cm
6km
sprint
Utero-tubal junction
Cervical mucins
100 X106
Sialic acid binding Ig Superfamily Lectins
Siglecs
Vaginal secretions
2
7 9
Swelling
modified from Hafner et al. 2013. Mucosal Immunology
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Functions of the Mammalian Sperm Glycome!
Cervical Mucins during cycle
and HA during pregnancy
Atomic Force Microscopy
Three gel-forming (MUC5B, MUC5AC, and MUC6) and two
transmembrane mucins (MUC16 and MUC1) were identified.
Tecle & Gagneux 2015 Mol. Repr. Develop.
Immune Modulation by the sperm glycocalyx!
Andersch-Björkman et al.,
Molecular & Cellular Proteomics (2007)
Brunelli et al.,
FASEB J.(2007)
Sperm hyaluronidase!
bisecting type N-glycans
Pang et al. JBC (2007)
Gamete glycocalyces
Theodore L. Tollner, Charles L. Bevins & Gary N. Cherr
Nature Reviews Urology 9, 365-375 (July 2012)
Egg Glycocalyx
•  Egg and Sperm have special glycoclyces.!
•  Egg Zone Pelucida!
Yauger, B, Boggs, NA, and Dean J.
Reproduction (2011)
Okabe, M Development (2013)
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Sialyl Lewis X on ZP N-glycans
UT Junction and Calmegin KO
Pang et al. Science (2011)
Nakanishi et al. Biol Reprod. (2004)!
UT Junction and ADAM3 KO
Egg Glycocalyx:
dynamic cortical granules
LCA, Lens culinaris agglutinin
ABL2 mAb
Liu et al. Reprod Biol Endocrinol. (2003)
Yamaguchi et al. Biol Reprod. (2009)!
A new view of mammalian fertilization
Okabe, M Asian J Andrology. (2015)
Okabe, M Development (2013)
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double fusion
Okabe, M Asian J Andrology. (2015)
Okabe, M Asian J Andrology. (2015)
Solid protein hand shake
Izumo-taisha!
出雲大社!
Japanese Shrine!
Juno!
Roman Goddess!
Bianchi, E. et al. Nature (2014)
With many Glycan Falvors!
Glycodelin Glycoforms
Immunological Paradox of Mammalian Fertilization!
Immunosuppressive
Anti-fertilizing
Seppälä et al. Human Repr. Update (2007)!
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Early stages of human development from fertilization to blastocyst formation
Glycan-mediated implantation!
Mouse Blastocyst!
stained for Neu5Gc!
sialic acid !
Making a home!
Ilekis. Potential placental molecular therapeutic targets. Am J Obstet Gynecol (2016)!
Vascular Remodelling!
Red-Horse et al. Development (2005)
Genbacev et al. Science (2003)!
Home Invasion!!
Red-Horse et al. Development (2005)
Anchoring and Invading!
Red-Horse et al. Development (2005)
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Syncytin Glycoprotein capture
Siglec 6 on Human Placenta
Brinkman-Van der Linden et al. Glycobiology. (2007)
Placentation
Brinkman-Van der Linden et al. Glycobiology. (2007)
Placentation preecclampsia
upregulation of Siglec 6
Preecclampsia: Pregnancy Associated Hypertension
Parham & Moffet Nature REV Immunol. (2013)
Placentation MUC16 as NK cell suppressor
MUC16 interacts with Siglec 9 on NK cells
Rumer et al. Reproductive Sciences. (2012)
Gubbels et al. Molecular Cancer (2010)
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Glycobiology of Human Reproduction - !
How Sugarcoating Solves Difficult Interactions!
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