Showing papers by "Worcester Foundation for Biomedical Research published in 2000"

Responses of olfactory receptor neurons in Utetheisa ornatrix to gender-specific odors.

Abstract: We recorded the electrophysiological responses of individual olfactory receptor neurons in sensilla trichodea on the antennae of adult arctiid moths, Utetheisa ornatrix, to stimulation with volatiles associated with both sexes. All trichoid sensilla contain at least two receptor neurons, each with distinct action potential amplitudes and waveforms, that respond dichotomously to male and female odors. Although, neither female neuron responds to extracts of coremata or the male-produced pheromone hydroxydanaidal, they do respond in a gender-specific manner to the volatiles emanating from whole pupae, hemolymph, thoracic froth, and adult animals of several ages. Thoracic froth, which contains pyrrolizidine alkaloids, is thought to play a role in defense. Froth from moths reared on diets, with or without added pyrrolizidine alkaloids, were equally effective in eliciting gender-specific patterns of response. Male trichoid receptor neurons respond to these same materials with similar patterns of activation. These receptor neurons provide information about substances, which we have termed "gender odors," that are persistently emitted by nearby animals. These substances do not appear to be the same as those already known to be involved in defense or the sexual dialog between individuals.

Detection of PACH1, a nuclear factor implicated in the transcriptional regulation of meiotic and early haploid stages of spermatogenesis.

Abstract: Spermatogenesis occurs in a series of well-defined stages and serves as an excellent model for lineage-specific cell development. Yet, little is known regarding the transcriptional mechanisms responsible for cell- and stage-dependent gene regulation in the male germ line. The rat and mouse proenkephalin genes are expressed from an alternative, spermatogenic cell-specific promoter specifically in meiotically-active pachytene spermatocytes and early post-meiotic spermatids. This promoter thus serves as an excellent model for defining transcriptional regulators involved in germ line-specific gene expression in meiotic cells. Previous transgenic studies identified a proximal, 51 bp 5'-flanking sequence containing two direct repeat elements that are absolutely required for in vivo proenkephalin promoter activity in spermatocytes and spermatids. Here, footprinting analyses were used to further delineate the specific interactions of a spermatogenic cell nuclear factor with the repeat elements within the proximal promoter region. This repeat-binding factor was also shown to be developmentally upregulated specifically in pachytene spermatocytes. Using Southwestern analysis, we have identified a unique nuclear protein enriched in pachytene spermatocytes that specifically recognizes the repeat elements within the proximal 5'-flanking sequence. We propose that this DNA binding factor, termed PACH 1, is a key transcriptional regulator of the proenkephalin and potentially other gene promoters, uniquely expressed during meiosis in the male germ line.

Activation of protein kinase C alters p34(cdc2) phosphorylation state and kinase activity in early sea urchin embryos by abolishing intracellular Ca2+ transients.

Abstract: The p34(cdc2) protein kinase, a universal regulator of mitosis, is controlled positively and negatively by phosphorylation, and by association with B-type mitotic cyclins. In addition, activation and inactivation of p34(cdc2) are induced by Ca(2+) and prevented by Ca(2+) chelators in permeabilized cells and cell-free systems. This suggests that intracellular Ca(2+) transients may play an important physiological role in the control of p34(cdc2) kinase activity. We have found that activators of protein kinase C can be used to block cell cycle-related alterations in intracellular Ca(2+) concentration ([Ca(2+)](i)) in early sea urchin embryos without altering the normal resting level of Ca(2+). We have used this finding to investigate whether [Ca(2+)](i) transients control p34(cdc2) kinase activity in living cells via a mechanism that involves cyclin B or the phosphorylation state of p34(cdc2). In the present study we show that the elimination of [Ca(2+)](i) transients during interphase blocks p34(cdc2) activation and entry into mitosis, while the elimination of mitotic [Ca(2+)](i) transients prevents p34(cdc2) inactivation and exit from mitosis. Moreover, we find that [Ca(2+)](i) transients are not required for the synthesis of cyclin B, its binding to p34(cdc2) or its destruction during anaphase. However, in the absence of interphase [Ca(2+)](i) transients p34(cdc2) does not undergo the tyrosine dephosphorylation that is required for activation, and in the absence of mitotic [Ca(2+)](i) transients p34(cdc2) does not undergo threonine dephosphorylation that is normally associated with inactivation. These results provide evidence that intracellular [Ca(2+)](i) transients trigger the dephosphorylation of p34(cdc2) at key regulatory sites, thereby controlling the timing of mitosis entry and exit.