PECULIARITIES OF THE STRUCTURE AND EXPRESSION OF HUMAN SPHINGOMYELIN SYNTHASE 1 GENE (SGMS1).

The article is a review of the results of the study of the structural and functional organization of the human sphingomyelin synthase 1 gene (SGMS1) in Human Molecular Genetics Department of Institute of Molecular Genetics RAS. SGMS1 gene encodes an essential enzyme which is involved in the synthesis of sphingomyelin and diacylglycerol from phosphatidylcholine and ceramide, wich determines its participation in the regulation of intracellular vesicular transport, cholesterol metabolism, cell proliferation, apoptosis and other significant processes. Our research has shown that the SGMS1 gene is located on the chromosome 10, has a size of 320 kb and contains more than 20 exons. A detailed study of the SGMS1 gene's structure allowed us to identify the variety of its transcripts. mRNA isoforms with different fragments of 5R untranslated region (5R UTR) and encoding the full length protein, as well as transcripts resulting from alternative combinations of exons and containing the coding region of the gene and 3R UTR have been discovered. We have found new transcripts among the products of SGMS1 gene ­ circular RNAs, which mostly contained sequences of multi-exon 5R UTR of the gene. They are conservative and predominantly expressed in the brain. Circular RNAs of SGMS1 gene had a large number of binding sites for a microRNA that may determine the functional significance of these molecules. The review describes the latest information about the structural and functional organization of the human gene SGMS1 as well as the features of its expression.

[PREPARATION OF HUMAN TISSUE PROTEIN EXTRACTS ENRICHED WITH THE SPHINGOMYELIN SYNTHASE 1].

Sphingomyelin synthase 1 (SMS 1) catalyzes sphingomyelin biosynthesis in eukaryotic cells. We previously studied the structure of the human SGMS1 gene, which encodes the enzyme and its numerous transcripts. The tissue-specific expression of the transcripts was also described. Analysis of the SMS1 protein expression in human tissues using immunoblotting of tissue extracts prepared in the RIPA (Radio Immuno-Precipitation Assay) buffer revealed a weak signal in renal cortex, testis, lung, and no signal in placenta and lymphatic node. In this work, a new method of preparation of the tissue protein extracts enriched with SMS1 was suggested. The method based on the consecutive extraction with a buffer containing 0.05 and 1 mg/ml of the Quillaja saponaria saponin allowed SMS1 to be detected in all tissues tested. The SMS1 content in the saponin extract of kidney cortex is about 12-fold higher compared to the RIPA extraction procedure.

[Alternative promoters localised in SGMS1 gene introns take part in regulation of its expression in human tissues].

Sphingomyelin synthase 1 (SMS1) is an enzyme of vital importance which is responsible for the synthesis of sphingomyelin and diacylglycerol from phosphatidylcholine and ceramide in eukaryotic cells. Previously we have investigated in detail the structure of SGMS1 human gene and identified a lot of its transcripts. We revealed the isoforms of mRNA differing in the 5'-UTR and coding the full-length protein, and also the transcripts arising from alternative combination of the exons localized in the coding region of the gene and 3'-UTR. From the results of computer analysis it follows that the synthesis of transcripts differing in the 5'-UTR is enabled by the different promoters of SGMS1 gene. It has been found in the present work by the method of real-time PCR that the content of five alternative transcripts of this gene, differing in the 5'-UTR, is substantially dissimilar among human tissues. In all the investigated tissues those transcripts are presented most prominently whose synthesis takes place under the control of the distal promoter including exon 1. In lesser extent are presented the transcripts including 5'-end exons whose synthesis is enabled by the promoters localized in introns of this gene. The differential level of content of SGMS1 gene transcripts, differing in the 5'-UTR, indicates that the use of the alternative promoters is tissue-specific and apparently strictly regulated. The structural organization of 5'-UTR variants of SGMS1 transcripts, directed by alternative promoters, is substantially different; this can provide regulation of the gene functioning on post-transcriptional level.