Expression of Tas1 Taste Receptors in Mammalian Spermatozoa: Functional Role of Tas1r1 in Regulating Basal Ca2+ and cAMP Concentrations in Spermatozoa

Background: During their transit through the female genital tract, sperm have to recognize and discriminate numerous chemical compounds. However, our current knowledge of the molecular identity of appropriate chemosensory receptor proteins in sperm is still rudimentary. Considering that members of the Tas1r family of taste receptors are able to discriminate between a broad diversity of hydrophilic chemosensory substances, the expression of taste receptors in mammalian spermatozoa was examined. Methodology/Principal Findings: The present manuscript documents that Tas1r1 and Tas1r3, which form the functional receptor for monosodium glutamate (umami) in taste buds on the tongue, are expressed in murine and human spermatozoa, where their localization is restricted to distinct segments of the flagellum and the acrosomal cap of the sperm head. Employing a Tas1r1-deficient mCherry reporter mouse strain, we found that Tas1r1 gene deletion resulted in spermatogenic abnormalities. In addition, a significant increase in spontaneous acrosomal reaction was observed in Tas1r1 null mutant sperm whereas acrosomal secretion triggered by isolated zona pellucida or the Ca2+ ionophore A23187 was not different from wild-type spermatozoa. Remarkably, cytosolic Ca2+ levels in freshly isolated Tas1r1-deficient sperm were significantly higher compared to wild-type cells. Moreover, a significantly higher basal cAMP concentration was detected in freshly isolated Tas1r1-deficient epididymal spermatozoa, whereas upon inhibition of phosphodiesterase or sperm capacitation, the amount of cAMP was not different between both genotypes. Conclusions/Significance: Since Ca2+ and cAMP control fundamental processes during the sequential process of fertilization, we propose that the identified taste receptors and coupled signaling cascades keep sperm in a chronically quiescent state until they arrive in the vicinity of the egg - either by constitutive receptor activity and/or by tonic receptor activation by gradients of diverse chemical compounds in different compartments of the female reproductive tract

Dual role of Miro protein clusters in mitochondrial cristae organisation and ER-Mitochondria Contact Sites

Mitochondrial Rho (Miro) GTPases localize to the outer mitochondrial membrane and are essential machinery for the regulated trafficking of mitochondria to defined subcellular locations. However, their sub-mitochondrial localization and relationship with other critical mitochondrial complexes remains poorly understood. Here, using super-resolution fluorescence microscopy, we report that Miro proteins form nanometer-sized clusters along the mitochondrial outer membrane in association with the Mitochondrial Contact Site and Cristae Organizing System (MICOS). Using knockout mouse embryonic fibroblasts (MEF) we show that Miro1 and Miro2 are required for normal mitochondrial cristae architecture and endoplasmic reticulum-mitochondria contacts sites (ERMCS). Further, we show that Miro couples MICOS to TRAK motor protein adaptors to ensure the concerted transport of the two mitochondrial membranes and the correct distribution of cristae on the mitochondrial membrane. The Miro nanoscale organization, association with MICOS complex and regulation of ERMCS reveal new levels of control of the Miro GTPases on mitochondrial functionality

Expression of inositol 1,4,5-trisphosphate receptors in mouse oocytes and early embryos: The type I isoform is upregulated in oocytes and downregulated after fertilization

A fertilization-induced increase in intracellular Ca2+ is responsible for initiating all of the events of egg activation. In mammals, the Ca2+ increase takes the form of a series of Ca2+ oscillations showing complex temporal and spatial properties. To understand the nature of these changes, we have investigated the expression patterns of the three isoforms of the inositol trisphosphate receptor (InsP(3)R) during oocyte maturation and preimplantation development. We find that mouse oocytes express mRNAs for all three InsP(3)R subtypes. Semiquantitative ratio reverse-transcriptase polymerase chain reaction shows that the type II isoform is the predominant message in mature oocytes, representing 67% of the InsP(3)R mRNA. In contrast, protein analysis reveals that the type I isoform accounts for all of the detectable InsP(3)R protein, despite representing only 20% of the InsP(3)R mRNA. The levels of InsP(3)R protein were examined to determine whether they correlated with the Ca2+ signaling events surrounding the fertilization process. Type I InsP(3)R protein increased during oocyte maturation and, in addition, within 8 h of fertilization underwent a dramatic decrease. During development to the blastocyst the level of type I InsP(3)R protein did not return to prefertilization levels and types II and III remained below our detection limit. The decrease in InsP(3)R protein after fertilization was found to correlate with a decrease in the sensitivity of InsP(3)-induced Ca2+ release. These studies show that the expression of InsP(3)R mRNA is developmentally regulated, that Ca2+ signaling at fertilization is mediated exclusively through the type I InsP(3)R, and that the InsP(3)R is downregulated after fertilization. (C) 1998 Academic Press