[Novel bidirectional promoter from human genome].

In human and other mammalian genomes a number of closely linked gene pairs transcribed in opposite directions are found. According to bioinformatic analysis up to 10% of human genes are arranged in this way. In present work the fragment of human genome was cloned that separates genes localized at 2p13.1 and oriented "head-to-head", coding for hypothetical proteins with unknown functions--CCDC (Coiled Coil Domain Containing) 142 and TTC (TetraTricopeptide repeat Containing) 31. Intergenic CCDC142-TTC31 region overlaps with CpG-island and contains a number of potential binding sites for transcription factors. This fragment functions as bidirectional promoter in the system ofluciferase reporter gene expression upon transfection of human embryonic kidney (HEK293) cells. The vectors containing genes of two fluorescent proteins--green (EGFP) and red (DsRed2) in opposite orientations separated by the fragment of CCDC142-TTC31 intergenic region were constructed. In HEK293 cells transfected with these vectors simultaneous expression of two fluorescent proteins is observed. Truncated versions of intergenic region were obtained and their promoter activity measured. Minimal promoter fragment contains elements Inr, BRE, DPE characteristic for TATA-less promoters. Thus, from the human genome the novel bidirectional promoter was cloned that can be used for simultaneous constitutive expression of two genes in human cells.

[Clinical and molecular genetic verification of Frasers syndrome, false male hermaphroditism and chronic glomerulonephritis (the first description in Russia)].

The Wilms tumor gene (WT1) encodes a transcription factor that plays a key role in the laying and differentiation of the kidneys and gonads. Mutations of the WT1 gene were detected in patients with the WAGR complex (Wilms tumor, aniridia, urogenital pathology, mental retardation), Denis-Drach syndrome (early renal failure, diffuse mesangial sclerosis, varying degrees of gonadal dysgenesis, high risk of Wilms tumor) and Fraser syndrome. The latter is characterized by a fully female phenotype with karyotype 46XY, focal segmental glomerulosclerosis with the development of renal failure in the 2nd decade of life, gonads in the form of cords and a high risk of gonadoblastoma. The presence of a heterozygous point mutation, which alters the donor site of splicing of the intron 9 of the WT1 gene, is also typical of Fraser syndrome. We present a case of characteristic clinical manifestations of Fraser syndrome in a patient in whom the diagnosis was confirmed by the detection of a mutation in the WT1 gene.

[Lipoid adrenal hyperplasia caused by earlier unknown 21STAR gene mutations].

Lipoid adrenal hyperplasia (LAH) is a most severe type of congenital adrenal cortical dysfunction (CACD). In this type of CACD, there is defect in the conversion of cholesterol to pregnenolone, as a consequence the production of all steroid hormones was impaired in both the adrenals and gonads. Defects of the STAR gene encoding for a steroidogenic acute regulatory (StAR) protein underlie the disease in most cases. Until the present time, there have been no reports on cases of LAH in the Russian literature. The diagnosis of LAH was established by the authors in three genetic girls aged 2.3 years, 6 and 7 months who had a normal structure of the external genitalia and in whom the disease was marked by manifestations of primary hypoadrenocorticism at the age of 21, 2, and 10 days, respectively. A hormonal study failed to show elevated levels of Cortisol and 17-hydroxyprogesterone in response to adrenocorticotropic hormone stimulation. A molecular genetic study revealed the following STAR gene defects: P129AC/W250X, IVS5-1G, and W147X, respectively. Р129ЛС, IVSS, and W147X mutations have not been earlier described. The data of the observation emphasizes the need of including LAH into the algorithm of differential diagnosis of CACD in children with the normal female structure of the external genitalia. The molecular genetic analysis should be prominent in diagnosing LAH, by taking into account the lack of hormonal markers of the disease.

[Fine structure, expression and polymorphism of the human growth hormone receptor gene]. / Tonkaia struktura, ékspressiia i polimorfizm gena retseptora gormona rosta cheloveka.

The human growth hormone receptor gene (GHR) is an example of complex transcription units. The gene has a very long 5'-regulatory region and contains multiple alternative 5'-untranslated exons that are spliced to the protein-coding exons. Its transcription is driven by several promoters that reside far upstream of the coding region. The complete nucleotide sequence of the human GHR gene is known. This paper summarizes the data on the human GHR gene structure analysis and mapping of alternative 5'-untranslated exons. The data of the promoter activity assay in the reporter gene transient expression system, as well as the results of studying the potential regulatory role of the short upstream open reading frame located in one of the 5'-untranslated exons are presented. The search for repeated elements in the human GHR gene was done, and several polymorphic markers that can be used in studies of GHR association with various diseases were characterized.

[Sex dimorphism in expression of alternative variants of mRNA for the growth hormone receptor in rat liver]. / Polovoi dimorfizm ékspressii al'ternativnykh variantov mRNK retseptora gormona rosta v pecheni krysy.

Two alternative variants of 5'-untranslated sequence of the rat growth hormone receptor (GHR) mRNA were previously described. These variants of mRNA appear to be produced by the splicing of primary transcripts initiated from alternative promoters. We employed the ribonuclease protection assay to reveal these two variants of the GHR mRNA in RNA preparations from the liver of male, normal female, and pregnant female rats. All tissues examined contain approximately the same amount of the GHR mRNA with type II 5'-untranslated sequence. To the contrary, the content of type I variant shows very pronounced sexual differences; its level is very low in the male liver, approximately equal to that of the type II variant in the female liver, and further increases in the pregnant female liver. These data suggest that the higher content of the GHR observed in the female liver and its elevation during gestation result from the accumulation of "female specific" type I variants of the GHR mRNA.

[Use of growth hormone genes for the diagnosis of the disease of dwarfism]. / Ispol'zovanie genov gormona rosta dlia diagnostiki zabolevaniia karlikovost'iu.

The pattern of BamHI fragments of DNA from three children suggested to suffer the isolated growth hormone deficiency type. IA was not different from normal pattern registered in blot hybridization with [32P]cDNA of the growth hormone gene. The data permits one to exclude the above mentioned disease that is characterized by the deletion of HGH-N gene. The analogous DNA restriction analysis using HindIII restriction endonuclease has shown, that neither the sick children, nor their parents carry the deletion in heterozygotic state. The study of normal polymorphism of the restriction fragments length has shown that as for as the frequency of polymorphic MspI restriction endonuclease sites A and B in the growth hormone gene cluster (0.67 and 0.75 respectively) is concerned the Russian population in Moscow is closer to Mediterranean one than to North-european.

[Genetic engineering of peptide hormones]. / Geneticheskaia inzheneriia peptidnykh gormonov.

The application of different approaches for preparing DNAs coding for peptide hormones was demonstrated. The libraries of human, bovine and porcine pituitaries cDNA were obtained starting from their total mRNAs. Screening of these libraries revealed clones containing human, bovine and porcine growth hormone sequences, cDNAs for bovine ACTH-beta-lipotropin precursor and for bovine and porcine prolactin. The gene of human calcitonin was created by combination of chemical and enzymatic synthesis. This synthetic gene was further cloned in pBR322. The expression of cloned human growth hormone cDNA under control of different Escherichia coli promoters was studied and physico-chemical and biological properties of the growth hormone produced by E. coli were tested.

[Comparison of the isoacceptor makeup of transfer RNA for 2 early stages in the development of loach embryos]. / Sopostavlenie izoaktseptornogo sostava transportnykh RNK dlia dvukh rannikh stadi56I RAZVITIIIA ZARODYSHEI V'IUNA.

The chromatographic profiles of isoacceptor tRNA's for 9 amino acids from the loach embryos at two early developmental stages were compared by the method of chromatography on sepharose column in the decreasing gradient of ammonium sulphate concentration to elucidate the possible role of tRNA in the control of early embryogenesis processes. Certain differences both in the proportions of some isoacceptor peaks and their relative positions on the chromatographic profile were found for 8 aminoacyl tRNA. Methionyl-tRNA was represented by one constant peak.