Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein.

Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Research Abstract Details 

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Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Abstract Text:

Computer-assisted database analysis of sequences homologous to human acid ceramidase (ASAH) revealed a 1233-bp cDNA (previously designated cPj-LTR) whose 266-amino-acid open reading frame had approximately 36% identity with the ASAH polypeptide. Based on this high degree of homology, we undertook further molecular characterization of cPj-LTR and now report the full-length cDNA sequence, complete gene structure (renamed human ASAHL since it is a human acid ceramidase-like sequence), chromosomal location, primer extension and promoter analysis, and transient expression results. The full-length human ASAHL cDNA was 1825 bp and contained an open-reading frame encoding a 359-amino-acid polypeptide that was 33% identical and 69% similar to the ASAH polypeptide over its entire length. Numerous short regions of complete identity were observed between these two sequences and two sequences obtained from the Caenorhabditis elegans genome database. The 30-kb human ASAHL genomic sequence contained 11 exons, which ranged in size from 26 to 671 bp, and 10 introns, which ranged from 150 bp to 6.4 kb. The gene was localized to the chromosomal region 4q21.1 by fluorescence in situ hybridization analysis. Northern blotting experiments revealed a major 2.0-kb ASAHL transcript that was expressed at high levels in the liver and kidney, but at relatively low levels in other tissues such as the lung, heart, and brain. Sequence analysis of the 5'-flanking region of the human ASAHL gene revealed a putative promoter region that lacked a TATA box and was GC rich, typical features of a housekeeping gene promoter, as well as several tissue-specific and/or hormone-induced transcription regulatory sites. 5'-Deletion analysis localized the promoter activity to a 1. 1-kb fragment within this region. A major transcription start site also was located 72 bp upstream from the ATG translation initiation site by primer extension analysis. Expression analysis of a green fluorescence protein/ASAHL fusion protein in COS-1 cells revealed a punctate, perinuclear distribution, although no acid ceramidase activity was detected in the transfected cells using a fluorescence-based in vitro assay system.

Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Publishing Authors By Initials

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Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Journal Published:

PUBLICATION TYPE: Research Support, U.S. Gov't,

Journal: Genomics

VOLUME: 62

Page Numbers: 232-41

Journal Abbreviation: Genomics

ISSN: 0888-7543

DAY: 1

MONTH: Dec

YEAR: 1999

Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Information

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LANGUAGE: eng

NlmUniqueID: 8800135

Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Keywords Mesh Terms:

KEYWORDS: Transfection

MESH TERMS: genetics

Chemical & Substance for Abstract: Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein. Information

Substance Name: NAAA protein, human

Registry Number: EC 3.5.-

Grant and Affiliation Information for Molecular cloning and characterization of a human cDNA and gene encoding a novel acid ceramidase-like protein.

AFFILIATION: Department of Human Genetics, Mount Sinai School of Medicine, New York 10029, USA.

Country: UNITED STATES

AGENCY: United States NICHD

GRANT: HD 28607

ACRONYM: HD

MEDLINETA: Genomics

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