Gonadotropini, receptori i regulacija

POLOŽAJ HIPOFIZE I VEZA SA HYPOT

Veza sa HPT i funkcija

HPT-AHYP portni sistem

RAZVOJ HIPOFIZE

RAZVOJ ĆELIJA

ADENOHIPOFIZE

RAZVOJ ĆELIJA

ADENOHIPOFIZE

RAZVOJ ĆELIJA

ADENOHIPOFIZE

HORMONI HPT-HYP

ĆELIJE A-HYP

Mouse pituitary tissue grown from embryonic stem cells. Fully grown glands produced hormones when transplanted into mice.

Nature 2011: 480 (57)

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Nature 2011: 480 (57)

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Nature 2011: 480 (57)

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Nature 2011: 480 (57) INFORMATIVNO!!!

Nature 2011: 480 (57)

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Nature 2011: 480 (57)

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Nature 2011: 480 (57)

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Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

GLOBALNI Ca2+ DOGAĐAJI

GLOBALNI Ca2+ DOGAĐAJI

Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

Ca2+ u regulaciji homeostaze i funkcije gonadotrofa i laktotrofa

GPCRs modulacija aktivnosti kanala u ćelijama adenohipofize

Spontana električna aktivnost i Ca2+ gonadotrofa i laktotrofa

Uloga GPCRs-ADCY na aktivnost membranskih kanala u ćelijama adenohipofize

Uloga receptora koji mobilišu Ca2+ i na aktivnost kanala i Ca2+ signaling u ćelijama adenohipofize

Spontana i receptorima regulisana električna aktivnost elija adenohipofize

Protein scaffolds in the coupling of synaptic exocytosis and endocytosis Volker Haucke, Erwin Neher & Stephan J. Sigrist Nature Reviews Neuroscience 12, 127-138 (March 2011)

Uloga Ca2+ zavisne egzocitoze u sekretornojaktivnosti gonadotrofa i laktotrofa

Ca2+ regulisana egzocitoza i proteini koji kao skele “povezuju” (coupling) egzocitozu i endocitozu

Endokrine i folikulostelatne ćelije adenohipofize

Folikulostelatne ćelije (FSc) adenohipofize

Intercelularni Ca2+ talasi u FSc

ADENOHIPOFIZA GONADOTROFI

Regulatcija sekrecije LH i FSH gondotrofima Release of luteinising hormone (LH) and follicle-stimulating hormone (FSH) is regulated by gonadotropin-releasing hormone (GnRH), which acts through the GnRH receptor (GnRH-R) to stimulate the formation of inositol 1,4,5-trisphosphate (InsP3). The latter then induces a cytosolic oscillator that releases a periodic burst of Ca2+ from the endoplasmic reticulum (ER). See text for further details. Berridge, M.J. (2012) Cell Signalling Biology; doi:10.1042/csb0001010

GONADOTROF

Oscilacije Ca2+ u gonadotrofima

GONADOTROPNI PROTEINI

a-GSU

DIFFERENT RESPONSE to GnRH frequencies

a-GSU – preferentially transcribed at high GnRH pulse frequencies LHB – at intermediate GnRH pulse frequencies FSHB – at lower GnRH pulse frequency

Biased signaling (“pristrasan” signaling)

GLIKOZILACIJA GONADOTROPINA

Veliki broj elemenata reguliše ekspresiju a-GSU

AR i GnRHR koriste iste regulatorne sekvence za obezbeđivanje tzv “moment-to-moment” regulacije a-GSU

Regulacije transkripcije a-GSU zavisi od aktivacije AR i GnRHR

Sekvenca hLHR i hFSHR (“sequence alignment”)

Interakcija između gonadotropnih receptora i hormona

Mesta “prirodno-nastalih” polimorfizama, “los-of-function” mutacija i “gain-of-function” mutacija u hLHR

Mesta “prirodno-nastalih” polimorfizama, “los-of-function” mutacija i “gain-of-function” mutacija u hFSHR

Sumirana regulacija transkripcije FSHR/Fshr

Organizacija FSHR/Fshr i LHCGR/Lhcgr i “genomsko okruženje”

Desenzitivizacija i “down”-regulacija kao mehanizmi modulacije odgovora receptora za gonadotropine

2012

Uloga glikozilacije u signalingu od receptora za gonadotropine

Veoma važna uloga glikozilacije u signalingu od receptora za gonadotropine

TRI MODELA AKTIVACIJE GONADOTROPINSKIH RECEPTORA

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NEKI PRIMERI SIGNALINGA OD AKTIVIRANIH GONADOTROPINSKIH RECEPTORA

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NEKI PRIMERI SIGNALINGA OD AKTIVIRANIH GONADOTROPINSKIH RECEPTORA

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NEKI PRIMERI SIGNALINGA GONADOTROPINSKIH RECEPTORA

NEKI PRIMERI SIGNALINGA GONADOTROPINSKIH RECEPTORA

NEKI PRIMERI SIGNALINGA GONADOTROPINSKIH RECEPTORA

REGULACIJA AKTIVNOSTI GONADOTROFA

REGULACIJA AKTIVNOSTI GONADOTROFA

REGULACIJA AKTIVNOSTI GONADOTROFA

REGULACIJA AKTIVNOSTI GONADOTROFA GnRH GnRI Androgeni Estrogeni Progesteron Aktivini Inhibini Folistatin PACAP cAMP/cGMP FOXO DMP1 ...

REGULACIJA AKTIVNOSTI GONADOTROFA

REGULACIJA AKTIVNOSTI GONADOTROFA

GnRH Signals Through Calcineurin, NFAT and JUN to Regulate Expression of at Least Three Genes Required for Gonadotrope Function BIOLOGY OF REPRODUCTION (2010)

Calcineurin is required for the GnRH effect on the activation of the FSHβ gene.

Pnueli L et al. Mol. Cell. Biol. 2011;31:5023-5036

REGULACIJA AKTIVNOSTI GONADOTROFA

Calcineurin is required for the GnRH effect on the activation of the FSHβ gene. (A) LβT2 cells were transfected with mFSH-luc, and in some cells CnA was overexpressed before treatment with GnRH and/or CsA, after which luciferase assays were performed. Data are presented as above, except that means are shown relative to the same treatment group (FSH, +CsA, or +CnA). n = 4. Pairwise analysis compared means within the same treatment group for cells with or without GnRH (NS, P > 0.05; **, P < 0.01; ***, P < 0.001). (B) LβT2 cells were either left untreated or treated with GnRH for as long as 60 min, and ChIP was carried out using an Ab against Pol II, endogenous NFAT3 (left), or FLAG (after overexpression of FLAG-tagged NFAT) (right). The presence of the Nur77 promoter was assessed by PCR. Also shown is amplification from the input samples and rabbit IgG controls.

(C) RT-PCR was carried out in LβT2 cells following transfection of siRNA to NFAT3, NFAT4, or GFP (as a control), with or without GnRH treatment, before amplification of a 380-bp fragment of the Nur77 coding sequence.

(D) Luciferase assays were carried out in LβT2 cells after transfection of FSHβ-luc either with siNFAT3 or with siGFP (300 ng) as a control. Data are presented as before. n = 4.

(E) Quantitative real-time PCR was used to measure endogenous levels of FSH or Nur77 mRNA in LβT2 cells after transfection of siNFAT3 (right) or siGFP as a control (left). Levels are expressed relative to those in siGFP-transfected, GnRH-treated cells. Means ± standard errors of the means (n = 3) are shown.

(F) LβT2 cells were transfected with FSHβ-luc together with mTORC1 and/or Nur77, and some of the cells were treated with GnRH. Luciferase assays were carried out, and data are presented as above. n = 4.

(G) LβT2 cells were transfected with FLAG-TORC1 and were either left untreated or exposed to GnRH for 4 h. IP was then carried out using an Ab against FLAG, and the precipitated proteins were analyzed by Western blot analysis using an Ab against TORC1 or Nur77.camb

REGULACIJA AKTIVNOSTI GONADOTROFA GnRH GnRI Aktivini Inhibini Folistatini cAMP/cGMP PACAP DMP1...

REGULACIJA ESKPRESIJA GONADOTROPINA

AKTIVINI I INHIBINI

Aktivin/Folistatin signaling

Aktivin/Folistatin signaling

REGULACIJA AKTIVNOSTI GONADOTROFA

Aktivin/Inhibin signaling

Aktivin/Inhibin signaling

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Uloga gonadotropina u formiranju modela interakcije transkripcionih faktora na promotoru gena za inhibin

Efekti FSH i aktivina u ovarijumu

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Aktivin/FSH stimulisana proliferacija

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Efekti aktivina i inhibina u testisu

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Efekti aktivina i inhibina u ovarijumu

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Šematski prikaz interakcije PACAP i GnRH u regulaciji ekspresije gena za gonadotropine

Pituitary Adenylate Cyclase Activating Peptide

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Regulaciji ekspresije gena za gonadotropine FOXO proteinima

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Uloga GnIH u regulaciji gonadotropina i njihovih receptora

Šematski prikaz hGnRIR (GPR147)

Uloga GnIH u regulaciji gonadotropina i njihovih receptora

Female-specific induction of rat pituitary dentin matrix protein-1 by GnRH. Kucka M et al. Mol Endocrinol. 2013 Nov;27(11):1840-1855

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Female-specific induction of rat pituitary dentin matrix protein-1 by GnRH. Kucka M et al. Mol Endocrinol. 2013 Nov;27(11):1840-1855

Hypothalamic GnRH is the primary regulator of reproduction in vertebrates, acting via the G protein-coupled GnRH receptor (GnRHR) in pituitary gonadotrophs to control synthesis and release of gonadotropins. To identify elements of the GnRHR-coupled gene network, GnRH was applied in a pulsatile manner for 6 hours to a mixed population of perifused pituitary cells from cycling females, mRNA was extracted, and RNA sequencing analysis was performed. This revealed 83 candidate-regulated genes, including a large number coding for secreted proteins. Most notably, GnRH induces a greater than 600-fold increase in expression of dentin matrix protein-1 (Dmp1), one of five members of the small integrin-binding ligand N-linked glycoprotein gene family. The Dmp1 response is mediated by the GnRHR, not elicited by other hypothalamic releasing factors, and is approximately 20-fold smaller in adult male pituitary cells. The sex-dependent Dmp1 response is established during the peripubertal period and independent of the developmental pattern of Gnrhr expression. In vitro, GnRH-induced expression of this gene is coupled with release of DMP1 in extracellular medium through the regulated secretory pathway. In vivo, pituitary Dmp1 expression in identified gonadotrophs is elevated after ovulation. Cell signaling studies revealed that the GnRH induction of Dmp1 is mediated by the protein kinase C signaling pathway and reflects opposing roles of ERK1/2 and p38 MAPK; in addition, the response is facilitated by progesterone. These results establish that DMP1 is a novel secretory protein of female rat gonadotrophs, the synthesis and release of which are controlled by the hypothalamus through the GnRHR signaling pathway. This advance raises intriguing questions about the intrapituitary and downstream effects of this new player in GnRH signaling.

INFORMATIVNO!!! Nešto od “doprinos”-a članova RES grupe istraživanju gonadotropa

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