The Role Of Mater In Cytoplasmic Lattice Formation, Endoplasmic Reticulum Distribution, And Calcium Homeostasis In Mouse Oocytes
Ca2+ oscillation is hallmark of mammalian fertilization that is thought to mediate egg activation process, and dramatic targeting of the endoplasmic reticulum (ER) to the microvillar subcortex, is known to provide calcium pools for subsequent activation and embryo development. While the mechanisms by which Ca2+ oscillation govern later embryonic development are poorly understood, emerging evidences indicate the critical role of Ca2+ oscillation in embryonic development, for example, by regulating recruitment of maternal mRNAs. Mater (Nlrp5) and Padi6 are oocyte-and early embryo-restricted maternal effect genes that are required for embryonic development beyond the 2-cell stage. Interestingly, PADI6 localizes to, and is required for the formation of oocyte-specific structure termed cytoplasmic lattices (CPLs). Based on similarity between MATER and PADI6, we hypothesized that MATER is also required for CPL formation. Indeed, ultrastructural analysis of Matertm/tm oocytes show ~90% reduced volume of CPLs and immuno-EM demonstrates the localization of MATER to CPLs, indicating MATER as a component of CPLs. Interestingly, Tubulin was also identified as a CPL component and that Tubulin interacts with PADI6. Given that Tubulin drives organelle movement, the role of Tubulin-PADI6-CPLs in organelle distribution during oocyte maturation was demonstrated. Given above observations, we further hypothesized that Mater plays a role in ER distribution and function, Ca2+ homeostasis in oocytes. We first investigated ER distribution by microinjecting DiI into oocytes and found that, as opposed to WT oocytes, the ER in metaphase-II Matertm/tm oocytes failed to concentrate around the spindle and cortical clusters were reduced. As cortical clusters are thought to be important for Ca2+ oscillation, then we tested if the pattern of Ca2+ oscillation is altered in Matertm/tm oocytes after in vitro fertilization. Intriguingly, Matertm/tm oocytes show altered Ca2+ oscillation pattern, lower amplitude and higher frequency. Then we demonstrated the reduced amount of intracellular Ca2+ stores in Matertm/tm oocytes, in part, contribute to such phenotype. Finally, the localization of IP3R-I, an essential player in releasing Ca2+ from ER, was altered in supporting of ER distribution. Taken together, this observation support the hypothesis that MATER is required for CPL formation and is involved in ER distribution and Ca2+ homeostasis in oocytes.
MATER; cytoplasmic lattice; calcium homeostasis
Coonrod, Scott A.
Schimenti, John C.; Parrish, Colin Ross; Clark, Theodore G.
Ph.D. of Veterinary Medicine
Doctor of Philosophy
dissertation or thesis