Family of serine hydrolase 1(FSH1). Previously: Domain of unknown function (DUF341). Contains human-OVCA2 Human Candidate tumor suppressor in ovarian cancer and yeast-FSH3 regulated by the Crt1 transcription factor, an effector of the DNA damage checkpoint pathway in Saccharomyces cerevisiae, may be involved in folate metabolism. Contains also the gene (dfr1) encoding FSH1 linked to a domain corresponding to dihydrofolate reductase (DHFR) in Schizosaccharomyces pombe. Fsh1p was identified as a lysophospholipase that specifically acts on lysophosphatidylserine (LPS) and impacts the lipid homeostasis in S. cerevisiae. Contains also Aspergillus terreus esterase LovG, which catalyzes the release of covalently bound dihydromonacolin L from LovB during lovastatin biosynthesis. OVCA2 has a strong preference for long-chain alkyl ester substrates (>10-carbons) and high selectivity against a variety of short, branched, and substituted esters
OBJECTIVES: To elucidate the role of FSH1 (family of serine hydrolase) in lipid homeostasis. RESULTS: Proteins in various species containing alpha/beta hydrolase domain are known to be involved in lipid metabolism. In silico analysis of the FSH1 gene in Saccharomyces cerevisiae revealed the presence of alpha/beta hydrolase domain (ABHD) and a lipase motif (GXSXG), however its function in lipid metabolism remained elusive. The overexpression of FSH1 in WT and fsh1delta cells showed a significant reduction in the cellular phospholipid levels and an increase in the triacylglycerol levels and lipid droplet (LD) number. Furthermore, the purified recombinant protein Fsh1p was identified as a lysophospholipase that specifically acts on lysophosphatidylserine (LPS) and impacts the lipid homeostasis in S. cerevisiae. CONCLUSIONS: These results depicted that Fsh1p has a role on lipid homeostasis and is a lysophospholipase that hydrolyzes lysophosphatidylserine (LPS).
        
Title: Comparative analysis of the human serine hydrolase OVCA2 to the model serine hydrolase homolog FSH1 from S. cerevisiae Bun JS, Slack MD, Schemenauer DE, Johnson RJ Ref: PLoS ONE, 15:e0230166, 2020 : PubMed
Over 100 metabolic serine hydrolases are present in humans with confirmed functions in metabolism, immune response, and neurotransmission. Among potentially clinically-relevant but uncharacterized human serine hydrolases is OVCA2, a serine hydrolase that has been linked with a variety of cancer-related processes. Herein, we developed a heterologous expression system for OVCA2 and determined the comprehensive substrate specificity of OVCA2 against two ester substrate libraries. Based on this analysis, OVCA2 was confirmed as a serine hydrolase with a strong preference for long-chain alkyl ester substrates (>10-carbons) and high selectivity against a variety of short, branched, and substituted esters. Substitutional analysis was used to identify the catalytic residues of OVCA2 with a Ser117-His206-Asp179 classic catalytic triad. Comparison of the substrate specificity of OVCA2 to the model homologue FSH1 from Saccharomyces cerevisiae illustrated the tighter substrate selectivity of OVCA2, but their overlapping substrate preference for extended straight-chain alkyl esters. Conformation of the overlapping biochemical properties of OVCA2 and FSH1 was used to model structural information about OVCA2. Together our analysis provides detailed substrate specificity information about a previously, uncharacterized human serine hydrolase and begins to define the biological properties of OVCA2.
Family of Serine Hydrolases (FSH) members FSH1, FSH2 and FSH3 in Saccharomyces cerevisiae share conserved sequences with the human candidate tumor suppressor OVCA2. In this study, hydrogen peroxide (H2O2) exposure increased the expression of both mRNA and protein levels of FSH3 in wild-type (WT) yeast cells. The deletion of FSH3 improved the yeast growth rate under H2O2-induction as compared to WT control cells. The overexpression of FSH3 in WT yeast cells caused an apoptotic phenotype, including accumulation of reaction oxygen species, decreased cell viability and cell death. The double deletions fsh1Delta fsh2Delta, fsh1Delta fsh3Delta and fsh2Delta fsh3Delta displayed increased growth compared to WT cells. However, the overexpression of FSH3 effectively inhibited cell growth in all double deletions. Moreover, the overexpression of FSH3 in cells lacking NUC1 did not cause any growth defect in the presence or absence of H2O2. Our results suggest that FSH3 induced apoptosis of yeast in a NUC1 dependent manner.
OBJECTIVES: To elucidate the role of FSH1 (family of serine hydrolase) in lipid homeostasis. RESULTS: Proteins in various species containing alpha/beta hydrolase domain are known to be involved in lipid metabolism. In silico analysis of the FSH1 gene in Saccharomyces cerevisiae revealed the presence of alpha/beta hydrolase domain (ABHD) and a lipase motif (GXSXG), however its function in lipid metabolism remained elusive. The overexpression of FSH1 in WT and fsh1delta cells showed a significant reduction in the cellular phospholipid levels and an increase in the triacylglycerol levels and lipid droplet (LD) number. Furthermore, the purified recombinant protein Fsh1p was identified as a lysophospholipase that specifically acts on lysophosphatidylserine (LPS) and impacts the lipid homeostasis in S. cerevisiae. CONCLUSIONS: These results depicted that Fsh1p has a role on lipid homeostasis and is a lysophospholipase that hydrolyzes lysophosphatidylserine (LPS).
        
Title: Comparative analysis of the human serine hydrolase OVCA2 to the model serine hydrolase homolog FSH1 from S. cerevisiae Bun JS, Slack MD, Schemenauer DE, Johnson RJ Ref: PLoS ONE, 15:e0230166, 2020 : PubMed
Over 100 metabolic serine hydrolases are present in humans with confirmed functions in metabolism, immune response, and neurotransmission. Among potentially clinically-relevant but uncharacterized human serine hydrolases is OVCA2, a serine hydrolase that has been linked with a variety of cancer-related processes. Herein, we developed a heterologous expression system for OVCA2 and determined the comprehensive substrate specificity of OVCA2 against two ester substrate libraries. Based on this analysis, OVCA2 was confirmed as a serine hydrolase with a strong preference for long-chain alkyl ester substrates (>10-carbons) and high selectivity against a variety of short, branched, and substituted esters. Substitutional analysis was used to identify the catalytic residues of OVCA2 with a Ser117-His206-Asp179 classic catalytic triad. Comparison of the substrate specificity of OVCA2 to the model homologue FSH1 from Saccharomyces cerevisiae illustrated the tighter substrate selectivity of OVCA2, but their overlapping substrate preference for extended straight-chain alkyl esters. Conformation of the overlapping biochemical properties of OVCA2 and FSH1 was used to model structural information about OVCA2. Together our analysis provides detailed substrate specificity information about a previously, uncharacterized human serine hydrolase and begins to define the biological properties of OVCA2.
Family of Serine Hydrolases (FSH) members FSH1, FSH2 and FSH3 in Saccharomyces cerevisiae share conserved sequences with the human candidate tumor suppressor OVCA2. In this study, hydrogen peroxide (H2O2) exposure increased the expression of both mRNA and protein levels of FSH3 in wild-type (WT) yeast cells. The deletion of FSH3 improved the yeast growth rate under H2O2-induction as compared to WT control cells. The overexpression of FSH3 in WT yeast cells caused an apoptotic phenotype, including accumulation of reaction oxygen species, decreased cell viability and cell death. The double deletions fsh1Delta fsh2Delta, fsh1Delta fsh3Delta and fsh2Delta fsh3Delta displayed increased growth compared to WT cells. However, the overexpression of FSH3 effectively inhibited cell growth in all double deletions. Moreover, the overexpression of FSH3 in cells lacking NUC1 did not cause any growth defect in the presence or absence of H2O2. Our results suggest that FSH3 induced apoptosis of yeast in a NUC1 dependent manner.
        
Title: FSH1 overexpression triggers apoptosis in Saccharomyces cerevisiae Gowsalya R, Ravi C, Arul M, Nachiappan V Ref: Antonie Van Leeuwenhoek, 112:1775, 2019 : PubMed
FSH1 belongs to the family of serine hydrolases in yeast and is homologous to the human ovarian tumor suppressor gene (OVAC2). Our preliminary results showed that cells lacking Fsh1p exhibit an increase in cell growth, and a decrease in the expression of AIF1 and NUC1 (apoptosis responsive genes) when compared to the wild type cells. Growth inhibition of cells overexpressing FSH1 is due to induction of cell death associated with cell death markers typical of mammalian apoptosis namely DNA fragmentation, phosphatidylserine externalization, ROS accumulation, Cytochrome c release, and altered mitochondrial membrane potential. When wild type cells were overexpressed with FSH1 there was up regulation of AIF1 level when compared to control cells suggesting that overexpression of FSH1 regulated cell death in yeast.
        
Title: FSH1 regulates the phenotype and pathogenicity of the pathogenic dermatophyte Microsporum canis Zhang F, Tan C, Xu Y, Yang G Ref: Int J Mol Med, 44:2047, 2019 : PubMed
Microsporum canis (M. canis) is a common pathogen that causes tinea capitis and is present worldwide. The incidence of M. canis infection, particularly tinea capitis, has been increasing in China. In our previous studies, family of serine hydrolases 1 (FSH1) was identified as a potential virulence factor in tinea capitis infection caused by M. canis. To determine the function of this gene in M. canis, FSH1 was knocked down using doublestranded RNA interference mediated by Agrobacterium tumefaciens. Reverse transcriptionquantitative PCR analysis was used to confirm gene knockdown. Loss of FSH1 expression by RNAi resulted in a minor phenotype alteration, but M. canis pathogenicity in guinea pig cutaneous infection was decreased compared with the wildtype strain. To the best of our knowledge, the present study is the first to demonstrate that FSH1 is associated with macroconidia septa formation and is an important contributor to M. canis virulence. These findings may advance the understanding of the function of the FSH1 gene and provide a foundation for future studies on macroconidia septa formation and pathogenicity of M. canis.
        
Title: LovG: the thioesterase required for dihydromonacolin L release and lovastatin nonaketide synthase turnover in lovastatin biosynthesis Xu W, Chooi YH, Choi JW, Li S, Vederas JC, Da Silva NA, Tang Y Ref: Angew Chem Int Ed Engl, 52:6472, 2013 : PubMed
OVCA2 is a putative serine-hydrolase. Performing protein profiling in human tumour cell lines, OVCA2 was detected in DAOY medulloblastoma cells as a high abundance protein. The protein was unambiguously identified by 2D gel-electrophoresis and MALDI-MS and MS/MS, its presence was confirmed by western blotting. Immunohistochemistry revealed expression in medulloblastoma and predominantly in oligodendrocytes. Computational approaches predicted functional motifs and domains, interaction with apoptosis-related protein BAG and 3D structure. In addition to the presence of OVCA2 in medulloblastoma, it was furthermore detectable in three out of 10 human tumour cell-lines as a high abundance protein probably suggesting a role in the tumour biology.
Yhr049w/FSH1 was recently identified in a combined computational and experimental proteomics analysis for the detection of active serine hydrolases in yeast. This analysis suggested that FSH1 might be a serine-type hydrolase belonging to the broad functional alphabeta-hydrolase superfamily. In order to get insight into the molecular function of this gene, it was targeted in our yeast structural genomics project. The crystal structure of the protein confirms that it contains a Ser/His/Asp catalytic triad that is part of a minimal alpha/beta-hydrolase fold. The architecture of the putative active site and analogies with other protein structures suggest that FSH1 may be an esterase. This finding was further strengthened by the unexpected presence of a compound covalently bound to the catalytic serine in the active site. Apparently, the enzyme was trapped with a reactive compound during the purification process.
        
Title: Identification of new genes regulated by the Crt1 transcription factor, an effector of the DNA damage checkpoint pathway in Saccharomyces cerevisiae Zaim J, Speina E, Kierzek AM Ref: Journal of Biological Chemistry, 280:28, 2005 : PubMed
The Crt1 (RFX1) protein in Saccharomyces cerevisiae is an effector of the DNA damage checkpoint pathway. It recognizes a 13-bp cis-regulatory element in the 5'-untranslated region (5'-UTR) of the ribonucleotide reductase genes RNR2, RNR3, and RNR4; the HUG1 gene; and itself. We calculated the weight matrix representing the Crt1p binding site motif according to analysis of the 5'-UTR sequences of the genes that are under its regulation. We subsequently searched the 5'-UTR sequences of all the genes in the yeast genome for the occurrence of this motif. The motif was found in regulatory regions of 30 genes. A statistical analysis showed that it is unlikely that a random gene cluster contains the motif conserved as well as the Crt1p binding site. Analysis of microarray data provided supporting evidence for five putative Crt1p targets: FSH3, YLR345W, UBC5, NDE2, and NTH2. We used reverse transcription-PCR to compare the expression levels of these genes in wild-type and crt1Delta strains. Our results indicated that FSH3, YLR345W, and NTH2 are indeed under the regulation of Crt1p. Sequence analysis of the FSH3p indicated that this protein may be involved in folate metabolism either by carrying serine hydrolase activity required for the novel metabolic pathway involving dihydrofolate reductase (DHFR) or by directly interacting with the DHFR enzyme. We postulate that Crt1p may influence deoxyribonucleotide synthesis not only by regulating expression of the RNR genes but also by modulating DHFR activity. FSH3p shares significant sequence similarity with the product of the human tumor suppressor gene OVCA2. YLR345Wp and NTH2p are enzymes involved in the central metabolism under stress conditions.
Retinoids, the natural and synthetic derivatives of vitamin A, have been shown to regulate the growth and differentiation of a wide variety of cell types and consequently have enormous potential as chemotherapeutic agents. We have previously identified 2 genes, termed OVCA1 and OVCA2, which are located in a small region showing a high frequency of allelic loss in breast and ovarian tumors and share a common exon. Recent studies have suggested that expression of OVCA1 may be influenced by retinoids. Therefore, we analyzed the expression of OVCA1 and OVCA2 in cells in response to treatment with all-trans retinoic acid (RA) and N-(4-hydroxyphenyl)retinamide (4HPR), or under conditions of low serum and confluence, to determine further the roles of OVCA1 and OVCA2 in cell growth, apoptosis and differentiation. We show that OVCA2 mRNA and protein are ubiquitously expressed and that they are downregulated in the lung cancer cell line Calu-6 after treatment with RA and 4HPR. In addition, we observed that OVCA2 protein is proteolytically degraded in response to RA and 4HPR treatment in a time- and dose-dependent manner in the promyelocytic leukemia cell line HL60. In contrast, expression of the candidate tumor suppressor OVCA1 was not downregulated by these treatments. Furthermore, we demonstrate that OVCA2 is evolutionarily conserved and shows regional homology with dihydrofolate reductases (DHFRs), specifically with hydrolase folds found in alpha-beta hydrolases. Our results are in contrast to a previous report and show that OVCA2, not OVCA1 mRNA and protein, is downregulated in response to RA and 4HPR.
A second tumor suppressor locus on 17p that is distinct from TP53 has been identified in brain, breast, lung, and ovarian tumors. Using allelic loss mapping and positional cloning methods, we have recently identified two novel genes, which we refer to as OVCA1 and OVCA2, that map to 17p13.3. The two genes are ubiquitously expressed and encode proteins of 443 and 227 amino acids, respectively, with no known functional motifs. Sequence comparison of OVCA1 and OVCA2 revealed extensive sequence identity and similarity to hypothetical proteins from Saccharomyces cerevisiae, Caenorhabditis elegans, and Rattus species. Northern blot analysis reveals that OVCA1 and OVCA2 mRNA were expressed in normal surface epithelial cells of the ovary, but the level of this transcript is significantly reduced or is undetectable in 92% (11/12) of the ovarian tumors and tumor cell lines analyzed. The location, high degree of amino acid conservation, and reduced expression in ovarian tumors and tumor cell lines suggest that decreased expression of these two genes contributes to ovarian tumorigenesis and should be considered candidate tumor suppressor genes.