Lopes, John

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Professor, Department of Microbiology, College of Natural Sciences
Last Name
Lopes
First Name
John
Discipline
Microbiology
Expertise
Combinatorial regulation of gene expression by basic helix-loop-helix proteins
Evolution of membrane phospholipid composition in the Saccharomyces genus
Genomic analysis of PI synthesis in yeast
Introduction
The goal of the research in my lab is to understand the transcriptional regulation of phospholipid biosynthesis in yeast, and how phospholipid biosynthesis is coordinated with other biological processes. To address this goal, my lab is carrying on two inter-related projects. One project is designed to determine how phospholipid biosynthesis is coordinated with other biological processes via a set of transcription factors that belong to the basic helix-loop-helix (bHLH) family. Another project is focused on the regulationof the only essential phospholipid biosynthetic gene ( PIS1 ) required for the synthesis of phosphatidylinositol (PI). This project is driven by the fact that virtually nothing is known about the transcriptional regulation of the PIS1 gene except that it is not coordinated with expression of the other phospholipid biosynthetic genes. We have been very successful with both projects publishing 38 papers with 2 more accepted for publication. My contributions in these areas are perhaps best evidenced by 4 solicited reviews. To carry out these projects, I have maintained a steady stream of extramural funding from the American Cancer Society and the National Science Foundation.
More recently, we have begun to examine the evolutionary conservation of the regulatory pathways that control phospholipid biosynthesis in the Saccharomyces genus. This project is in its infancy and I plan to apply for funding in the near future. Each of these three projects is summarized below.
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1995 - 20002
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    PublicationOpen Access
    The promoter of the yeast INO4 regulatory gene:a model of the simplest yeast promoter
    (2000) Lopes, John; Robinson, Kelly
    In Saccharomyces cerevisiae, the phospholipid biosynthetic genes are transcriptionally regulated in response to inositol and choline. This regulation requires the transcriptional activator proteins Ino4p and Ino2p, which form a heterodimer that binds to the UASINO element. We have previously shown that the promoters of the INO4 and INO2 genes are among the weakest promoters characterized in yeast. Because little is known about the promoters of weakly expressed yeast genes, we report here the analysis of the constitutive INO4 promoter. Promoter deletion constructs scanning 1,000 bp upstream of theINO4 gene identified a small region (−58 to −46) that is absolutely required for expression. S1 nuclease mapping shows that this region contains the transcription start sites for the INO4gene. An additional element (−114 to −86) modestly enhancesINO4 promoter activity (fivefold). Thus, the region required for INO4 transcription is limited to 68 bp. These studies also found that INO4 gene expression is not autoregulated by Ino2p and Ino4p, despite the presence of a putative UASINO element in the INO4promoter. We further report that the INO4 steady-state transcript levels and Ino4p levels are regulated twofold in response to inositol and choline, suggesting a posttranscriptional mechanism of regulation.
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    PublicationOpen Access
    Autoregulated expression of the yeast INO2 and INO4 helix-loop-helix genes effects cooperative regulation on their target genes
    (1995) Lopes, John; Ashburner, B.
    In the yeast Saccharomyces cerevisiae, the phospholipid biosynthetic genes are highly regulated at the transcriptional level in response to the phospholipid precursors inositol and choline. In the absence of inositol and choline (derepressing), the products of the INO2 and INO4 genes form a heteromeric complex which binds to a 10-bp element, upstream activation sequence INO (UASINO), in the promoters of the phospholipid biosynthetic genes to activate their transcription. In the presence of inositol and choline (repressing), the product of the OPI1 gene represses transcription dictated by the UASINO element. Curiously, we identified a UASINO-like element in the promoters of both the INO2 and INO4 genes. The presence of the UASINO element in these two promoters suggested that the mechanism for the inositol-choline response would involved regulating expression of the two activator genes. Using a cat reporter gene, we find that INO2-cat expression was regulated 12-fold in response to inositol and choline but that INO4-cat was constitutively expressed. We further observed that INO2-cat was not expressed in either an ino2 or an ino4 mutant strain and was constitutively overexpressed in an opi1 mutant strain. Expression of the INO4-cat gene was affected only by mutation in the INO4 gene itself. Therefore, INO2-cat transcription is regulated by the products of both the INO2 and INO4 genes whereas INO4 must interact with another protein to activate its own transcription. Our data show that derepression of phospholipid biosynthetic gene expression involves two mechanisms: increasing the levels of the INO2 and INO4 gene products and inactivating the OPI1-mediated repression mechanism. We propose a model suggesting that this dual mechanism of regulation accounts for the observed cooperative stimulation of IN01 and CH01 gene expression (phospholipids biosynthetic genes).