Professor and Chair
Department of Cell Biology

Contact

Phone: 860-679-2661
Email: ljaffe@uchc.edu
Office: E6032

UConn School of Medicine
263 Farmington Avenue
Farmington, CT 06030

Curriculum Vitae

Research Interests

Research in the Jaffe lab concerns the physiological mechanisms that regulate the oocyte cell cycle and fertilization. Currently, our studies are focused on regulation of meiosis in mammalian ovarian follicles by cyclic nucleotides.

Meiosis prepares the oocyte for fertilization, by reducing the number of copies of each gene from two to one, such that the female and male genomes from oocytes (eggs) and sperm can combine to make a new individual. In mammalian oocytes, meiosis begins during embryonic development of the mother, and then arrests in prophase for a prolonged period. Much of the process of meiosis occurs within a spherical complex of somatic cells, called granulosa cells, that make up a follicle; the outer granulosa cells are called mural granulosa, and the granulosa cells closest to the oocyte are called cumulus (see photo below). During each reproductive cycle, a group of follicles grow to a stage at which luteinizing hormone from the pituitary can act on the mural granulosa cells, to cause resumption of meiosis and ovulation of a mature fertilizable egg.

The follicle functions as a coordinated system in which processes in the granulosa cells, as well as processes in the oocyte itself, regulate meiotic progression in the oocyte.

Work in the Jaffe lab has established that a G_s G-protein and an associated receptor, GPR3, both located in the oocyte, contribute to maintaining meiotic prophase arrest in mouse oocytes (Mehlmann et al., 2002; 2004). The activity of the G_s G-protein leads to production of cyclic AMP in the oocyte, and high cAMP keeps the cell cycle arrested during storage in the ovary .

We have also determined that the granulosa cells of the follicle contribute to maintaining meiotic prophase arrest by regulating the hydrolysis of cAMP in the oocyte. This regulation involves another cyclic nucleotide, cyclic GMP, which diffuses from the granulosa cells into the oocyte, where it inhibits a cAMP phosphodiesterase, PDE3A, and thus maintains the high cAMP that maintains meiotic arrest (Norris et al., 2009). Then in response to luteinizing hormone (LH), which acts on a G-protein coupled receptor in the mural granulosa cells, the signaling system in the follicle switches, such that cAMP in the oocyte decreases and meiosis proceeds. This occurs primarily because LH signaling lowers cGMP in the granulosa cells (Norris et al., 2009).

The cGMP decrease in the granulosa cells begins within less than a minute after LH application (Shuhaibar et al., 2015). We can monitor this change using confocal microscopy of follicles from mice expressing a FRET sensor for cGMP, cGi500. The cGMP sensor mice were made by the lab of our collaborator, Robert Feil (University of Tubingen).The LH-induced cGMP decrease occurs in a wave progressing inwards. In the outer granulosa cells, where the LH receptors are located (Baena, et al., 2020), cGMP decreases to half of its plateau value within ~3 minutes. cGMP in the oocyte falls to half its plateau value within ~10 minutes, as a result of diffusion into the large volume of the surrounding granulosa cells. Supporting this diffusion-based mechanism for LH control of oocyte cGMP, we have previously shown that a 326-Da tracer, comparable in size to cGMP, diffuses from the oocyte through gap junctions to the granulosa cells with similar kinetics (Norris et al., 2008).

Cyclic GMP is produced in the mural granulosa and cumulus cells by the guanylyl cyclase natriuretic peptide receptor 2 (NPR2), which is activated by C-type natriuretic peptide (CNP); CNP is produced by the mural granulosa cells (work from the lab of John Eppig ; see Zhang et al., 2010, Science 330, 366-369). Our work, in collaboration with the lab of Lincoln Potter (University of Minnesota), has shown that LH signaling reduces cGMP in part by reducing its production, through rapid dephosphorylation and inactivation of the guanylyl cyclase NPR2 and a slower reduction of its agonist CNP (Robinson et al., 2012; Egbert et al., 2014; Shuhaibar et al, 2016). LH exposure also results in rapid phosphorylation of the phosphodiesterase PDE5. This is accompanied by a rapid increase in PDE5 activity, indicating that in parallel with decreasing cGMP production, LH signaling increases cGMP hydrolysis (Egbert et., 2014; Egbert et al., 2016, 2018).

Working model of how LH signaling rapidly decreases cGMP in the mural granulosa cells, and then via cGMP diffusion through gap junctions, decreases cGMP in the oocyte, leading to meiotic resumption. Before LH exposure, cGMP concentrations are elevated throughout the follicle, due to a high rate of production of cGMP by the NPR2 guanylyl cyclase in the mural granulosa and cumulus cells. cGMP phosphodiesterases, including PDE1 and PDE5, hydrolyze cGMP at a rate equal to its production, thus keeping the cGMP concentration at a constant level. Through gap junctions that connect all cells of the follicle, cGMP diffuses into the oocyte, where it inhibits the activity of PDE3A, maintaining cAMP at a level that inhibits meiotic resumption. The cAMP in the oocyte is produced by an adenylyl cyclase in the oocyte that is kept active by the constitutive activity of the Gs-coupled receptor GPR3. When LH binds to its receptor in the mural granulosa cells, the activation of Gs and possibly other G-proteins results in dephosphorylation of NPR2, which decreases its rate of production of cGMP. Activation of the LH receptor also increases phosphorylation of PDE5, which increases its rate of hydrolysis of cGMP. Due to reduced NPR2 activity and increased cGMP phosphodiesterase activity, the concentration of cGMP in the mural granulosa cells decreases. Through the series of gap junctions that connects the oocyte to the large volume of the mural granulosa cells, cGMP in the oocyte diffuses down its concentration gradient, and the resulting decrease in oocyte cGMP relieves the inhibition of PDE3A in the oocyte, such that cAMP decreases. This model depicts only events occuring in the first 20 minutes after LH exposure. Subsequent events, including a decrease in gap junction permeability, an increase in EGF receptor ligands, and a decrease in C-type natriuretic peptide, also contribute to maintaining cGMP at the low level that triggers meiotic resumption. Modified from Shuhaibar et al., 2015.

Common mechanisms of cGMP signaling in ovarian follicles and in the growth plate of bone.

Using the concepts and methods that we developed for our studies of LH signaling in ovarian follicles, we discovered that a similar signaling pathway contributes to FGF-induced inhibition of bone growth (Shuhaibar et al., 2017, 2021). In both ovary and bone, hormonal signaling results in dephosphorylation and inactivation of NPR2, thus decreasing cGMP; in the ovary, this causes meiotic resumption, while in bone, it causes a decrease in growth. Thus the basic mechanisms that we discovered in the ovary are significant in other physiological systems as well. See Leia  Shuhaibar's web page. Uconn today.

 

Currently, we are investigating the possibility that the same phosphatase might mediate NPR2 dephosphorylation in both ovarian follicles and chondrocytes.   Parallel studies of the two systems, using phosphomass spectroscopy, are providing clues  (collaboration with Ivan Silbern and Henning Urlaub, Max Planck Institute, Gottingen); see Jeremy Egbert's web page.

Lab Members

 

Movie: Microinjection of an antral follicle-enclosed mouse oocyte

Selected Publications

Shuhaibar, L.C.,  Kaci, N., Egbert, J.R., Horville, T., Loisay, L., Vigone, G., Uliasz, T.F., Dambroise,E., Swingle, M.R.,  Honkanen, R.E., Duplan, M.B., Jaffe, L.A., Legeai-Mallet, L. (2021) Phosphatase inhibition by LB-100 enhances BMN-111 stimulation of bone growth..pdf JCI insight. 6 (9):141426.

Baena, V., Owen, C.M., Uliasz, T.F., Lowther, K.M., Yee, S.-P., Terasaski, M., Egbert, J.E., and Jaffe, L.A. (2020). Cellular heterogeneity of the LH receptor and its significance for cyclic GMP signaling in mouse preovulatory follicles. Endocrinology  161 (7):bqaa074.

Egbert, J.R., Fahey, P.G., Reimer, J., Owen, C.M, Evsikov, A.V., Nikolaev, V.O., Griesbeck, O., Ray, R.S., Tolias, A.S., and Jaffe, L.A. (2019) Follicle-stimulating hormone and luteinizing hormone increase Ca2+ in the granulosa cells of mouse ovarian follicles. Biol. Reprod., 101(2), 433–444.

Egbert, J.R., Yee, S.-P., and Jaffe, L.A. (2018). Luteinizing hormone signaling phosphorylates and activates the cyclic GMP phosphodiesterase PDE5 in mouse ovarian follicles, contributing an additional component to the hormonally induced decrease in cyclic GMP that reinitiates meiosis. Develop. Biol. 435:6-14.

Shuhaibar, L.C., Robinson, J.W., Vigone, G., Shuhaibar, N.P., Egbert, J.R., Baena, V., Uliasz, T.V., Kaback, D., Yee, S.-P., Feil, R., Fisher, M.C., Dealy, C.N., Potter, L.R., and Jaffe, LA. (2017). Dephosphorylation of the NPR2 guanylyl cyclase contributes to inhibition of bone growth by fibroblast growth factor. eLife 6:e31343.

Robinson, J.W., Egbert, J.R., Davydova, J., Schmidt, H., Jaffe, L.A., and Potter, L.R. (2017). Dephosphorylation is the mechanism of fibroblast growth factor inhibition of guanylyl cyclase-B. Cellular Signaling 40:222-229.

Jaffe LA and Egbert JR, (2017) Regulation of Mammalian Oocyte Meiosis by Intercellular Communication Within the Ovarian Follicle. Annu. Rev. Physiol. 79: 237-260.

Egbert J.R., Uliasz T.F., Shuhaibar, L.C., Geerts A., Wunder F., Kleiman R.J., Humphrey J.M., Lampe P.D., Artemyev, N.O., Rybalkin, S.D., Beavo, J.A., Movsesian, M.A., and Jaffe, L.A. (2016) Luteinizing hormone causes phosphorylation and activation of the cyclic GMP phosphodiesterase PDE5 in rat ovarian follicles, contributing, together with PDE1 activity, to the resumption of meiosis. Biol. Reprod. 94(5):110. Supplement

Shuhaibar, L.C., Egbert, J.R., Edmund, A.B., Uliasz, T.F., Dickey, D.M., Yee, S.P., Potter, L.R., and Jaffe, L.A. (2016). Dephosphorylation of juxtamembrane serines and threonines of the NPR2 guanylyl cyclase is required for rapid resumption of oocyte meiosis in response to luteinizing hormone. Dev Biol. 409:194-201. Supplement

Shuhaibar, L.C., Egbert, J.R., Norris, R.P., Lampe, P.D., Nikolaev, V.O., Thunemann, M., Wen, L., Feil, R., and Jaffe, L.A. (2015). Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles. Proc. Natl. Acad. Sci. USA 112:5527-5532.

Egbert, J.R., Shuhaibar, L.C., Edmund, A.B., Van Helden, D.A., Robinson, J.W., Uliasz, T.F., Baena, V., Geerts, A., Wunder, F., Potter, L.R., and Jaffe, L.A. (2014). Dephosphorylation and inactivation of the NPR2 guanylyl cyclase in the granulosa cells contributes to the LH-induced decrease in cGMP that causes meiotic resumption in rat oocytes. Development. 141:3594-3604.

Robinson, J.W., Zhang, M., Shuhaibar, L.C., Norris, R.P., Geerts, A., Wunder, F., Eppig, J.J., Potter, L.R., and Jaffe, L.A. (2012). Luteinizing hormone reduces the activity of the NPR2 guanylyl cyclase in mouse ovarian follicles, contributing to the cyclic GMP decrease that promotes resumption of meiosis in oocytes. Dev. Biol. 366: 308–316. Supplement

Norris, R.P., Freudzon, M., Nikolaev, V.O., and Jaffe, L.A. 2010. Epidermal growth factor receptor kinase activity is required for gap junction closure and for part of the decrease in ovarian follicle cGMP in response to LH. Reproduction140: 655-662.

Norris, R.P., Ratzan, W.J., Freudzon, M., Mehlmann, L.M., Krall, J., Movsesian, M.A., Wang, H., Ke, H., Nikolaev, V.O., and Jaffe, L.A. (2009). Cyclic GMP from the surrounding somatic cells regulates cyclic AMP and meiosis in the mouse oocyte. Development 136: 1869-1878. Supplement

Jaffe, L.A., Norris, R.P., Freudzon, M., Ratzan, W.J., and Mehlmann, L.M. (2009). Microinjection of follicle-enclosed mouse oocytes. Methods Mol. Biol. 518:157-173.

Norris, R.P., Freudzon, M., Mehlmann, L.M., Cowan, A.E., Simon, A.M., Paul, D.L., Lampe, P.D., and Jaffe, L.A. (2008). Luteinizing hormone causes MAPK-dependent phosphorylation and closure of Cx43 gap junctions in mouse ovarian follicles: one of two paths to meiotic resumption. Development 135:3229-3238. Supplement

Norris, R.P., Freudzon, L., Freudzon, M., Hand, A.R., Mehlmann, L.M., and Jaffe, L.A. (2007). A Gs linked receptor maintains meiotic arrest in mouse oocytes, but luteinizing hormone does not cause meiotic resumption by terminating receptor Gs signaling. Develop. Biol. 310: 240-249.

Freudzon, L., Norris, R.P., Hand, A.R., Tanaka, S., Saeki, Y., Jones, T.L.Z., Rasenick, M.M., Berlot, C.H., Mehlmann, L.M., and Jaffe, L.A. 2005. Regulation of meiotic prophase arrest in mouse oocytes by GPR3, a constitutive activator of the Gs G protein. J. Cell Biol. 171: 255-265.

Jaffe, L.A., and Terasaki, M. 2004. Quantitative microinjection of oocytes, eggs and embryos. Meth. Cell Biol. 74: 219-242.

Kalinowski, R.R., Berlot, C.H., Jones, T.L.Z., Ross, L.F., Jaffe, L.A., and Mehlmann, L.M. 2004. Maintenance of meiotic prophase arrest in vertebrate oocytes by Gs protein-mediated pathway. Develop. Biol. 267: 1-13.

Mehlmann, L.M., Saeki, Y., Tanaka, S., Brennan, T.J., Evsikov, A.V., Pendola, F.L., Knowles, B.B., Eppig, J.J., and Jaffe, L.A. 2004. The Gs linked receptor GPR3 maintains meiotic arrest in mammalian oocytes. Science 306: 1947-1950. Supplement

Mehlmann, L.M., Jones,T.L.Z, and Jaffe, L.A. 2002. Meiotic arrest in the mouse follicle maintained by a Gs protein in the oocyte. Science 297: 1343-1345.

Jaffe, L.A. 1993. The experimental process: the electrical polyspermy block. Essay in the textbook Molecular and Cellular Biology, by Stephen L. Wolfe. Wadsworth Publishing Co., pp. 1110-1111.

Jaffe, L.A. 1976. Fast block to polyspermy in sea urchin eggs is electrically mediated. Nature 261:68‑71.

Research in the Jaffe lab is supported by a grant from the Eunice Kennedy Shriver National Institute of Child Health and Human Development (R37HD014939).