Gene_locus Report for: mouse-Ces1e

The mouse (Mus musculus) is the premier animal model for understanding human disease and development. Here we show that a comprehensive understanding of mouse biology is only possible with the availability of a finished, high-quality genome assembly. The finished clone-based assembly of the mouse strain C57BL/6J reported here has over 175,000 fewer gaps and over 139 Mb more of novel sequence, compared with the earlier MGSCv3 draft genome assembly. In a comprehensive analysis of this revised genome sequence, we are now able to define 20,210 protein-coding genes, over a thousand more than predicted in the human genome (19,042 genes). In addition, we identified 439 long, non-protein-coding RNAs with evidence for transcribed orthologs in human. We analyzed the complex and repetitive landscape of 267 Mb of sequence that was missing or misassembled in the previously published assembly, and we provide insights into the reasons for its resistance to sequencing and assembly by whole-genome shotgun approaches. Duplicated regions within newly assembled sequence tend to be of more recent ancestry than duplicates in the published draft, correcting our initial understanding of recent evolution on the mouse lineage. These duplicates appear to be largely composed of sequence regions containing transposable elements and duplicated protein-coding genes; of these, some may be fixed in the mouse population, but at least 40% of segmentally duplicated sequences are copy number variable even among laboratory mouse strains. Mouse lineage-specific regions contain 3,767 genes drawn mainly from rapidly-changing gene families associated with reproductive functions. The finished mouse genome assembly, therefore, greatly improves our understanding of rodent-specific biology and allows the delineation of ancestral biological functions that are shared with human from derived functions that are not.

Egasyn (esterase-22), a member of the nonspecific carboxylesterase multigene family (E.C. 3.1.1.1), is the endoplasmic reticulum (ER)-targeting protein of beta-glucuronidase. We utilized the polymerase chain reaction (PCR) in the eventual isolation of murine egasyn cDNAs. PCR primers were based upon: (1) partial amino acid sequences derived from egasyn peptides and (2) a conserved active site region shared by carboxylesterases. The amino acid sequence deduced from the PCR product matched that obtained from egasyn protein. This product was utilized as a probe to screen a cDNA library. Two cDNAs whose composite sequence encoded an open reading frame of 562 amino acids were isolated. A message size of 1700-2000 bp was revealed by RNA blot hybridization analysis. S1 nuclease protection analyses detected mRNA in liver, kidney, lung, and submandibular gland, but not in spleen, brain, and testes. Genetic mapping confirmed the location of an egasyn cDNA fragment in cluster 1 of the esterase region on chromosome 8. Transfection of COS cells with the 2022-bp cDNA resulted in the expression of esterase activity, which comigrated on native gels with liver esterase-22. The features of the deduced amino acid sequence of the egasyn cDNA are compared with previously characterized carboxylesterases and with other lumenal ER proteins.

We report biochemical, immunological, and genetic studies which demonstrate that an accessory protein with the essential features of mouse egasyn is complexed with and stabilizes a portion of beta-glucuronidase in microsomes of rat liver. The accessory protein exists as a complex with beta-glucuronidase since it coprecipitates with beta-glucuronidase after treatment of extracts with a specific beta-glucuronidase antibody. The two proteins are associated by noncovalent bonds since they are easily dissociated at elevated temperatures. Only 20-25% of total liver accessory protein is complexed with microsomal beta-glucuronidase. The remainder exists as a free form. The molecular weight of the accessory protein is 61 to 63 kDa depending upon the rat strain of origin. This protein, like mouse egasyn, has esterase catalytic activity and is concentrated in microsomes. The accessory protein is genetically polymorphic with at least four alleles. Combined biochemical and genetic evidence indicates it is identical with esterase-3 of the rat. Also, both mouse egasyn and rat esterase-3 react with antisera to egasyn and to rat esterase-3, indicating they are homologous proteins. Several inbred rat strains lack microsomal beta-glucuronidase. The same strains lack the accessory protein, suggesting that stabilization of beta-glucuronidase in rat microsomes requires egasyn.

The mammalian carboxylesterase 1 (Ces1/CES1) family comprises several enzymes that hydrolyze many xenobiotic chemicals and endogenous lipids. To investigate the pharmacological and physiological roles of Ces1/CES1, we generated Ces1 cluster knockout (Ces1 (-/-) ) mice, and a hepatic human CES1 transgenic model in the Ces1 (-/-) background (TgCES1). Ces1 (-/-) mice displayed profoundly decreased conversion of the anticancer prodrug irinotecan to SN-38 in plasma and tissues. TgCES1 mice exhibited enhanced metabolism of irinotecan to SN-38 in liver and kidney. Ces1 and hCES1 activity increased irinotecan toxicity, likely by enhancing the formation of pharmacodynamically active SN-38. Ces1 (-/-) mice also showed markedly increased capecitabine plasma exposure, which was moderately decreased in TgCES1 mice. Ces1 (-/-) mice were overweight with increased adipose tissue, white adipose tissue inflammation (in males), a higher lipid load in brown adipose tissue, and impaired blood glucose tolerance (in males). These phenotypes were mostly reversed in TgCES1 mice. TgCES1 mice displayed increased triglyceride secretion from liver to plasma, together with higher triglyceride levels in the male liver. These results indicate that the carboxylesterase 1 family plays essential roles in drug and lipid metabolism and detoxification. Ces1 (-/-) and TgCES1 mice will provide excellent tools for further study of the in vivo functions of Ces1/CES1 enzymes.

The mouse (Mus musculus) is the premier animal model for understanding human disease and development. Here we show that a comprehensive understanding of mouse biology is only possible with the availability of a finished, high-quality genome assembly. The finished clone-based assembly of the mouse strain C57BL/6J reported here has over 175,000 fewer gaps and over 139 Mb more of novel sequence, compared with the earlier MGSCv3 draft genome assembly. In a comprehensive analysis of this revised genome sequence, we are now able to define 20,210 protein-coding genes, over a thousand more than predicted in the human genome (19,042 genes). In addition, we identified 439 long, non-protein-coding RNAs with evidence for transcribed orthologs in human. We analyzed the complex and repetitive landscape of 267 Mb of sequence that was missing or misassembled in the previously published assembly, and we provide insights into the reasons for its resistance to sequencing and assembly by whole-genome shotgun approaches. Duplicated regions within newly assembled sequence tend to be of more recent ancestry than duplicates in the published draft, correcting our initial understanding of recent evolution on the mouse lineage. These duplicates appear to be largely composed of sequence regions containing transposable elements and duplicated protein-coding genes; of these, some may be fixed in the mouse population, but at least 40% of segmentally duplicated sequences are copy number variable even among laboratory mouse strains. Mouse lineage-specific regions contain 3,767 genes drawn mainly from rapidly-changing gene families associated with reproductive functions. The finished mouse genome assembly, therefore, greatly improves our understanding of rodent-specific biology and allows the delineation of ancestral biological functions that are shared with human from derived functions that are not.

Egasyn-beta-glucuronidase complex is located at the luminal site of liver microsomal endoplasmic reticulum. When organophosphorus insecticides (OP) are incorporated into the liver microsomes, they become tightly bound to egasyn, a carboxylesterase isozyme, and subsequently, beta-glucuronidase (BG) is dissociated and released into blood. Consequently, the increase in plasma BG activity becomes a good biomarker of OP exposure. Thus, the single administration of EPN (O-ethyl O-p-nitrophenylphenylphosphonothioate), acephate and chlorpyrifos increased plasma BG activity in approximately 100-fold the control level in rats. The increase in plasma BG activity after OP exposure is a much more sensitive biomarker of acute OP exposure than acetylcholinesterase (AChE) inhibition.

Egasyn is an accessory protein of beta-glucuronidase (beta-G) in the liver microsomes. Liver microsomal beta-G is stabilized within the luminal site of the microsomal vesicles by complexation with egasyn which is one of the carboxylesterase isozymes. We investigated the effects of organophosphorus compounds (OPs) such as insecticides on the dissociation of egasyn-beta-glucuronidase (EG) complex. The EG complex was easily dissociated by administration of OPs, i.e. fenitrothion, EPN, phenthionate, and bis-beta-nitrophenyl phosphate (BNPP), and resulting beta-G dissociated was released into blood, leading to the rapid and transient increase of plasma beta-G level with a concomitant decrease of liver microsomal beta-G level. In a case of phenthionate treatment, less increase in plasma beta-G level was observed, as compared with those of other OPs. This may be explained by the fact that phenthionate was easily hydrolyzed by carboxylesterase. Similarly, carbamate insecticides such as carbaryl caused rapid increase of plasma beta-G level. In contrast, no significant increase of plasma beta-G level was observed when pyrethroid insecticides were administered to rats. This is due to the fact that pyrethroids such as phenthrin and allethrin were easily hydrolyzed by A-esterase as well as carboxylesterase. On the other hand, addition of OPs to the incubation mixture containing liver microsomes caused the release of beta-G from microsomes to the medium. From these in vivo and in vitro data, it is concluded that increase of the plasma beta-G level after OP administration is much more sensitive biomarker than cholinesterase inhibition to acute intoxication of OPs and carbamates.

Egasyn (esterase-22), a member of the nonspecific carboxylesterase multigene family (E.C. 3.1.1.1), is the endoplasmic reticulum (ER)-targeting protein of beta-glucuronidase. We utilized the polymerase chain reaction (PCR) in the eventual isolation of murine egasyn cDNAs. PCR primers were based upon: (1) partial amino acid sequences derived from egasyn peptides and (2) a conserved active site region shared by carboxylesterases. The amino acid sequence deduced from the PCR product matched that obtained from egasyn protein. This product was utilized as a probe to screen a cDNA library. Two cDNAs whose composite sequence encoded an open reading frame of 562 amino acids were isolated. A message size of 1700-2000 bp was revealed by RNA blot hybridization analysis. S1 nuclease protection analyses detected mRNA in liver, kidney, lung, and submandibular gland, but not in spleen, brain, and testes. Genetic mapping confirmed the location of an egasyn cDNA fragment in cluster 1 of the esterase region on chromosome 8. Transfection of COS cells with the 2022-bp cDNA resulted in the expression of esterase activity, which comigrated on native gels with liver esterase-22. The features of the deduced amino acid sequence of the egasyn cDNA are compared with previously characterized carboxylesterases and with other lumenal ER proteins.

We report biochemical, immunological, and genetic studies which demonstrate that an accessory protein with the essential features of mouse egasyn is complexed with and stabilizes a portion of beta-glucuronidase in microsomes of rat liver. The accessory protein exists as a complex with beta-glucuronidase since it coprecipitates with beta-glucuronidase after treatment of extracts with a specific beta-glucuronidase antibody. The two proteins are associated by noncovalent bonds since they are easily dissociated at elevated temperatures. Only 20-25% of total liver accessory protein is complexed with microsomal beta-glucuronidase. The remainder exists as a free form. The molecular weight of the accessory protein is 61 to 63 kDa depending upon the rat strain of origin. This protein, like mouse egasyn, has esterase catalytic activity and is concentrated in microsomes. The accessory protein is genetically polymorphic with at least four alleles. Combined biochemical and genetic evidence indicates it is identical with esterase-3 of the rat. Also, both mouse egasyn and rat esterase-3 react with antisera to egasyn and to rat esterase-3, indicating they are homologous proteins. Several inbred rat strains lack microsomal beta-glucuronidase. The same strains lack the accessory protein, suggesting that stabilization of beta-glucuronidase in rat microsomes requires egasyn.