Store-operated calcium entry (SOCE) is the predominant Ca2؉ influx pathway in non-excitable cells and is activated in response to depletion of intracellular Ca 2؉ stores. We have studied SOCE regulation during Xenopus oocyte meiosis. SOCE can be measured readily in stage VI Xenopus oocytes arrested at the G 2 -M transition of the cell cycle, either by Ca 2؉ imaging or by recording the SOCE current. However, following meiotic maturation, SOCE can no longer be activated by store depletion. We have characterized the time course of SOCE inactivation during oocyte maturation, and show that SOCE inactivates almost completely, in a very short time period, at the germinal vesicle breakdown stage of meiosis. This acute inactivation offers an opportunity to better understand SOCE regulation.Ionic calcium (Ca 2ϩ ) is a universal second messenger important for many cellular responses ranging from gene expression to fertilization (1). Ca 2ϩ signaling is mediated by a rise in cytoplasmic Ca 2ϩ , either by Ca 2ϩ release from intracellular stores or Ca 2ϩ influx from the extracellular space. In nonexcitable cells the primary Ca 2ϩ influx pathway is store-operated calcium entry (SOCE) 1 (2). SOCE is activated in response to depletion of intracellular calcium stores and has been implicated in several cellular processes, including T-cell activation (3, 4) and regulation of exocytosis (5-7). However the mechanism(s) coupling store depletion to SOCE activation remain unknown. The following three models have been proposed: "physical coupling," "diffusible messenger," and "vesicle fusion." The physical coupling hypothesis proposes that SOCE channels couple directly to a Ca 2ϩ sensor on the endoplasmic reticulum membrane, possibly the IP 3 receptor, in analogy to the dihydropyridine-ryanodine receptor interaction in skeletal muscle (8 -10). Whereas the diffusible messenger hypothesis argues that store depletion results in the generation of a diffusible messenger that opens SOCE channels (11-13), the vesicle fusion model proposes that SOCE channels are inserted in the plasma membrane following store depletion (14). Although there is some evidence for each model (2), it is not yet clear what the coupling mechanism is and whether SOCE is activated by the same mechanism in different cell types.Despite the importance and ubiquitous nature of Ca 2ϩ signaling pathways, their role and regulation during the cell cycle remain controversial. Probably the clearest example of a Ca 2ϩ requirement during the cell cycle is at fertilization, where a Ca 2ϩ signal is necessary and sufficient for egg activation and the initiation of embryonic development (15, 16). Oocytes of both Xenopus and mammals are arrested at the G 2 -M transition of the cell cycle (15, 17). Before such oocytes become competent to be fertilized and able to support embryonic development, they undergo a maturation process called meiotic (or oocyte) maturation. During this maturation period oocytes enter meiosis, complete the first meiotic division with the extrusion of a polar body,...

Effective treatment regimens for pulmonary tuberculosis are difficult to assess because of the slow growth rate of Mycobacterium tuberculosis in culture and its protracted clearance from sputum. A rapid method that reflects effective antimicrobial activity would markedly advance evaluation of treatment and promote the assessment of new antituberculosis drugs. Conventional methods measure the progressive reduction of numbers of acid-fast bacilli in the sputum smear and the clearance of organisms in sputum culture. In this study, we measured levels of M. tuberculosis 85B (alpha antigen) messenger RNA (mRNA), 16S ribosomal RNA (rRNA), and IS6110 DNA in patients' sputa to ascertain whether they could serve as potential surrogate markers of response to chemotherapy. Sputum specimens were sequentially collected for up to a year from 19 smear-positive pulmonary tuberculosis patients receiving an optimal drug treatment regimen. Nucleic acids were isolated from these specimens, and two M. tuberculosis molecular targets (mRNA, DNA) were quantified, using the ABI Prism 7700 Sequence Detection System. The Mycobacterium genus-specific 16S rRNA was quantified with a limiting dilution RT-PCR assay. Results show that levels of 85B mRNA declined after initiation of therapy, as did viable M. tuberculosis colony counts, with 90% of patients becoming negative for both markers after 2 mo of treatment. The rapid disappearance of M. tuberculosis mRNA from sputum suggests that it is a good indicator of microbial viability and a useful marker for rapid assessment of response to chemotherapy.