[Model sharings by analyzing work flow and system construction].
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Efficiency, Organizational↗
Biomedical subjects
Hiroyuki Kubota
Publications and source records attributed to Hiroyuki Kubota.
The yeast eIF4E-associated protein Eap1p attenuates GCN4 translation upon TOR-inactivation.
Amino acid-starved yeast activates the eIF2alpha kinase Gcn2p to suppress general translation and to selectively derepress the transcription factor Gcn4p, which induces various biosynthetic genes to elicit general amino acid control (GAAC). Well-fed yeast activates the target of rapamycin (TOR) to stimulate translation via the eIF4F complex. A crosstalk was demonstrated between the pathways for GAAC and TOR signaling: the TOR-specific inhibitor rapamycin activates Gcn2p. Here we demonstrate that, upon TOR-inactivation, the putative TOR-regulated eIF4E-associated protein Eap1p likely functions downstream of Gcn2p to attenuate GCN4 translation via a mechanism independent of eIF4E-binding, thereby constituting another interface between the two pathways.
Amino Acid Sequence↗
Transcriptomic and proteomic analysis of a 14-3-3 gene-deficient yeast.
BMH1 and BMH2 encode Saccharomyces cerevisiae 14-3-3 homologues whose exact functions have remained unclear. The present work compares the transcriptomic and proteomic profiles of the wild type and a BMH1/2-deficient S. cerevisiae mutant (bmhDelta) using DNA microarrays and two-dimensional polyacrylamide gel electrophoresis. It is reported here that, although the global patterns of gene and protein expression are very similar between the two types of yeast cells, a subset of genes and proteins (a total of 220 genes) is significantly induced or reduced in the absence of Bmh1/2p. These genes include approximately 60 elements that could be linked to the reported phenotypes of the bmhDelta mutant (e.g., accumulation of glycogen and hypersensitivity to environmental stress) and/or could be the potential downstream targets of interacting partners of Bmh1/2p such as Msn2p and Rtg3p. Importantly, >30% of the identified genes (71 genes) were found to be associated with carbon (C) and nitrogen (N) metabolism and transport, thereby suggesting that Bmh1/2p may play a major role in the regulation of C/N-responsive cellular processes. This study presents the first comprehensive overview of the genes and proteins that are affected by the depletion of Bmh1/2p and extends the scope of knowledge of the regulatory roles of Bmh1/2p in S. cerevisiae.
14-3-3 Proteins↗
Rapamycin-induced translational derepression of GCN4 mRNA involves a novel mechanism for activation of the eIF2 alpha kinase GCN2.
When starved for amino acids, Saccharomyces cerevisiae accumulates uncharged tRNAs to activate its sole eukaryotic initiation factor (eIF) 2alpha kinase GCN2. Subsequent phosphorylation of eIF2alpha impedes general translation, but translationally derepresses the transcription factor GCN4, which induces expression of various biosynthetic genes to elicit general amino acid control response. By contrast, when supplied with enough nutrients, the yeast activates the target of rapamycin signaling pathway to stimulate translation initiation by facilitating the assembly of eIF4F. A cross-talk was suggested between the two pathways by rapamycin-induced translation of GCN4 mRNA. Here we show that rapamycin causes an increase in phosphorylated eIF2alpha to translationally derepress GCN4. This increment is not observed in the cells expressing mammalian non-GCN2 eIF2alpha kinases in place of GCN2. It is thus suggested that rapamycin does not inhibit dephosphorylation of eIF2alpha but rather activates the kinase GCN2. This activation seems to require an interaction between the kinase and uncharged tRNAs, because rapamycin, similar to amino acid starvation, fails to induce eIF2alpha phosphorylation in the cells with GCN2 defective in tRNA binding. However, in contrast with amino acid starvation, rapamycin activates GCN2 without increasing the amount of uncharged tRNAs, but presumably by modifying the tRNA binding affinity of GCN2.
Antifungal Agents↗
Spontaneous rupture of peripancreatic lymph node with hepatocellular carcinoma metastasis: report of an autopsy case with massive peritoneal bleeding.
Hepatocellular carcinoma (HCC) has a tendency for fatal spontaneous rupture leading to massive hemorrhage. The majority of such ruptures of HCC occur in the liver, while a few previous studies showed that such HCC rupture developed at metastatic sites including the lung, pleura, spleen, and peritoneum. We here report a case of hepatitis C virus-related HCC with spontaneous rupture of a peripancreatic lymph node with HCC metastases. A 61-year-old Japanese man died of hepatic failure after therapy for cirrhosis and HCC for 6 years. At autopsy, the liver showed diffuse multinodular HCC in both lobes. Metastases were found in lumbar vertebral bones and abdominal lymph nodes of the hepatic hilar, peripancreatic and perigastric regions. One lymph node (4 cm in diameter) around the pancreatic head was found ruptured, and blood coagula (100 g) was present around the ruptured node. Pure blood of 2000 ml was noted in the peritoneal cavity. Cirrhosis was not recognized. Histologically, the liver tumors and metastases in the lumbar vertebral bones and abdominal lymph nodes were poorly-differentiated HCC of Edmondson's grade III. The ruptured lymph node was almost replaced by poorly-differentiated HCC, and the ruptured site showed hemorrhage, breakdown of the capsule, and ischemic changes. Non-tumorous liver showed chronic hepatitis C. The immediate cause of death was suggested to be circulatory insufficiency due to rupture of the lymph node and/or hepatic failure. The present case showed that abdominal lymph nodes with HCC metastases may rupture and cause severe hemorrhage in the peritoneal cavity, leading to death.
Journal Article↗
Roles for the two-hybrid system in exploration of the yeast protein interactome.
Comprehensive analysis of protein-protein interactions is a challenging endeavor of functional proteomics and has been best explored in the budding yeast. The yeast protein interactome analysis was achieved first by using the yeast two-hybrid system in a proteome-wide scale and next by large-scale mass spectrometric analysis of affinity-purified protein complexes. While these interaction data have led to a number of novel findings and the emergence of a single huge network containing thousands of proteins, they suffer many false signals and fall short of grasping the entire interactome. Thus, continuous efforts are necessary in both bioinformatics and experimentation to fully exploit these data and to proceed another step forward to the goal. Computational tools to integrate existing biological knowledge buried in literature and various functional genomic data with the interactome data are required for biological interpretation of the huge protein interaction network. Novel experimental methods have to be developed to detect weak, transient interactions involving low abundance proteins as well as to obtain clues to the biological role for each interaction. Since the yeast two-hybrid system can be used for the mapping of the interaction domains and the isolation of interaction-defective mutants, it would serve as a technical basis for the latter purpose, thereby playing another important role in the next phase of protein interactome research.
Computational Biology↗