Biomedical subjects

M Fiszman

Publications and source records attributed to M Fiszman.

At least 19 recordsLinked to original sources

PubMed1997

Organization and sequence of human cardiac myosin binding protein C gene (MYBPC3) and identification of mutations predicted to produce truncated proteins in familial hypertrophic cardiomyopathy.

Cardiac myosin binding protein C (MyBP-C) is a sarcomeric protein belonging to the intracellular immunoglobulin superfamily. Its function is uncertain, but for a decade evidence has existed for both structural and regulatory roles. The gene encoding cardiac MyBP-C (MYBPC3) in humans is located on chromosome 11p11.2, and mutations have been identified in this gene in unrelated families with familial hypertrophic cardiomyopathy (FHC). Detailed characterization of the MYBPC3 gene is essential for studies on gene regulation, analysis of the role of MyBP-C in cardiac contraction through the use of recombinant DNA technology, and mutational analyses of FHC. The organization of human MYBPC3 and screening for mutations in a panel of French families with FHC were established using polymerase chain reaction, single-strand conformation polymorphism, and sequencing. The MYBPC3 gene comprises > 21,000 base pairs and contains 35 exons. Two exons are unusually small in size, 3 bp each. We found six new mutations associated with FHC in seven unrelated French families. Four of these mutations are predicted to produce truncated cardiac MyBP-C polypeptides. The two others should each produce two aberrant proteins, one truncated and one mutated. The present study provides the first organization and sequence for an MyBP-C gene. The mutations reported here and previously in MYBPC3 result in aberrant transcripts that are predicted to encode significantly truncated cardiac MyBP-C polypeptides. This spectrum of mutations differs from the ones previously observed in other disease genes causing FHC. Our data strengthen the functional importance of MyBP-C in the regulation of cardiac work and provide the basis for further studies.

Base Sequence↗

PubMed1995

Cardiac myosin binding protein-C gene splice acceptor site mutation is associated with familial hypertrophic cardiomyopathy.

Familial hypertrophic cardiomyopathy (FHC) is an autosomal dominant disease characterized by a ventricular hypertrophy predominantly affecting the interventricular septum and associated with a large extent of myocardial and myofibrillar disarray. It is the most common cause of sudden death in the young. In the four disease loci found, three genes have been identified which code for beta-myosin heavy chain, cardiac troponin T and alpha-tropomyosin. Recently the human cardiac myosin binding protein-C (MyBP-C) gene was mapped to chromosome 11p11.2 (ref. 8), making this gene a good candidate for the fourth locus, CMH4 (ref. 5). Indeed, MyBP-C is a substantial component of the myofibrils that interacts with several proteins of the thick filament of the sarcomere. In two unrelated French families linked to CMH4, we found a mutation in a splice acceptor site of the MyBP-C gene, which causes the skipping of the associated exon and could produce truncated cardiac MyBP-Cs. Mutations in the cardiac MyBP-C gene likely cause chromosome 11-linked hypertrophic cardiomyopathy, further supporting the hypothesis that hypertrophic cardiomyopathy results from mutations in genes encoding contractile proteins.

Amino Acid Sequence↗

PubMed1995

Inhibition of proliferation in 8-week-old mdx mouse muscle fibroblasts in vitro.

Our purpose is to understand why mdx muscle does not show the progressive degeneration observed in human Duchenne muscular dystrophy (DMD) muscle. In the mouse, the regenerative process compensates for the necrosis of the muscle fibers, particularly during the acute phase of the disease (5-9 weeks). In DMD muscle, there is a gradual failure of the regenerative process and the muscle fibers are replaced by connective and fatty tissue. We propose that distinct properties of mdx and DMD muscle fibroblasts could be one of the reasons for the differences between the mdx and DMD phenotypes. We found that fibroblasts taken from human DMD and control muscle had similar in vitro proliferative capacities. The proliferation rate of mouse muscle fibroblasts decreased during the acute phase of the disease, and inhibition was complete in fibroblasts from 8-week-old mdx mice. Moreover, the medium conditioned by these cells inhibited fibroblast proliferation. The effect was specific for fibroblasts, since this conditioned medium stimulated myoblast proliferation, as did control fibroblast-conditioned medium. These results suggest that 8-week-old mdx mouse muscle fibroblasts produce an inhibitor of their own proliferation and a growth factor specific for myoblasts in vitro. If these factors are secreted in vivo, the growth inhibitory factory may stop fibroblast proliferation whereas the mitogenic activity could stimulate satellite cell proliferation, thus favouring muscle regeneration.

Adolescent↗

PubMed1993

Muscle-specific expression of the acetylcholine receptor alpha-subunit gene requires both positive and negative interactions between myogenic factors, Sp1 and GBF factors.

The dependence of the muscle-specific enhancer of the acetylcholine receptor alpha-subunit gene on other domains of the promoter has been analysed by performing point mutagenesis and modular reconstitution of the enhancer--promoter sequences. The enhancer is inactive in the absence of the proximal region containing an Sp1 binding site and an overlapping G-C homopolymer binding factor site (referred to as GBF). The proximal region can be replaced by an Sp1 binding site from SV40 or an MEF-2 binding site from the muscle creatine kinase gene. Specific mutation of the Sp1 site markedly affects transactivation by CMD1 or myogenin. Mutation of the GBF binding site leads to higher promoter activity in primary cultures of chick myotubes or in quail fibroblasts. In addition, binding of a purified Sp1 protein prevents the binding of GBF in vitro. It is proposed that in the case of the alpha-subunit promoter, the myogenic factors activate transcription in cooperation with Sp1, and that GBF contributes to muscle-specific expression of the promoter by interfering with Sp1 binding in nonmuscle muscle cells or myoblasts.

Animals↗

PubMed1990

In vitro splicing of mutually exclusive exons from the chicken beta-tropomyosin gene: role of the branch point location and very long pyrimidine stretch.

The chicken beta-tropomyosin gene contains 11 exons, two of which are spliced into mRNA only in skeletal muscle. One pair of alternative exons, 6A and 6B, is found in the middle of the gene; they are spliced in a mutually exclusive manner. The non-muscle splice 6A-7 is by far the predominant in vitro reaction in a HeLa cell nuclear extract. A minor product is the 6A-6B splice, which is excluded in all tissues. This minor product results from the use of a branch point located 105 nt upstream of the 3' end of the intron separating exons 6A and 6B. The region between the branch point sequence and the final AG contains a stretch of approximately 80 pyrimidines. We have examined the role of the distance of the branchpoint to the 3' splice site and of the sequences between these two elements. Our results suggest that at least two cis-acting elements contribute to the mutual exclusivity of exons 6A and 6B. The intron between exons 6A and 6B is intrinsically poorly 'spliceable' both because the branch point is too far upstream of the 3' end of the intron to give efficient splicing and because of the particular sequence lying between this branch point and the 3' splice site.

Animals↗

PubMed1989

Multiple mRNAs encode peripherin, a neuronal intermediate filament protein.

Three cDNA clones of 1.6 (3u), 1.2 (5g) and 0.6 (5b) kbp, specific for peripherin, a neuronal intermediate filament protein (IFP), have been isolated from a murine neuroblastoma cell lambda gt11 library by immunoscreening using peripherin antiserum. Antibodies eluted from the fusion proteins produced by clones 3u and 5g recognize the peripherin spots on immunoblots. Where they overlap the three cDNAs have identical sequences. cDNA 5g exhibits the closest homology to type III IFP cDNAs. cDNA 3u is identical to the corresponding region of cDNA 5g, except for the insertion of a 96 bp fragment at a position corresponding to the junction of exons 4 and 5 in type III IFP cDNAs. cDNA 5b is also identical to the corresponding region of cDNA 5g, except for the deletion of a 62 bp fragment at the junction of exons 8 and 9 in type III IFP cDNAs. S1 mapping experiments performed with probes covering the 3' end of the two unexpected regions show that three distinct mRNAs correspond to the three cDNAs. Moreover, three peripherin products, two minor 61 and 56 kd products in addition to the major 58 kd peripherin, are observed when poly(A)+ RNA is in vitro translated, the 61 kd peripherin being translated from the 3u-selected RNA. The three RNAs originate from alternative splicing of a unique peripherin gene, thus generating polymorphism of peripherin.

Amino Acid Sequence↗

PubMed1985

Rous sarcoma virus SRC gene expression on the growth of quail embryo skin fibroblasts and the establishment of permanent cell lines.

Permanent cell lines of Quail embryo fibroblasts appear in cultures of cells infected with a wild type strain of Rous sarcoma virus (SR-RSV) or with its temperature sensitive transformation mutants (ts-T) (NYts68 and PA101) following a three step process. In step one, infected cells grow twice as fast as the control. The second step consists of a crisis during which the cell population is stationary for four to five weeks. Towards the fourth week several foci of cell growth are observed in the flasks. Respreading of the content of these flasks yields permanent lines. This constitutes the third step of the population evolution. In step one the growth rate of the infected cells is the same irrespective of the incubation temperature (36 degrees C or 41 degrees C) whereas the level of the pp60v-src activity is considerably depressed at 41 degrees C for NYts68 and PA101. Foci do not appear at restrictive temperature in the ts infected population and permanent lines are not recovered under that condition. These lines grow ony at 36 degrees C. It can be shown that the virus which they produce is not modified with respect to the temperature sensitivity of the src gene expression since newly infected fibroblasts grow equally well in step one at both 36 degrees C and 41 degrees C, and stop after the same number of generations. This finding suggests that the events which, during the crisis period, lead to the establishment of permanent lines, take place at the cellular level but depend on the activity of the pp60v-src protein for their occurrence or their expression.

Animals↗

PubMed1983

Fast and slow chicken skeletal muscles contain different alpha and beta tropomyosins.

Avian tropomyosin has been purified from fast skeletal muscles (breast muscle and posterior latissimus dorsi : PLD) and from a slow skeletal muscle (anterior latissimus dorsi : ALD) and the alpha and beta subunits have been further separated using preparative gel electrophoresis. These subunits have been subjected to partial proteolysis using different proteolytic enzymes. In this communication we show that this procedure allows to distinguish not only between fast and slow alpha tropomyosin but also between fast and slow beta tropomyosin. Furthermore we have raised an antiserum against the fast alpha tropomyosin and we present evidence to show that this antiserum does not cross-react with the slow alpha tropomyosin. These results are taken to indicate that all these tropomyosin subunits represent different gene products.

Animals↗

PubMed1983

[A thermosensitive characteristic of the Schmidt Rupin strain of Rous sarcoma virus affecting the longevity of infected lines].

Quail embryo skin fibroblasts infected with wild type Rous sarcoma virus SR-RSV grow, at 36 or 41 degrees C, twice as fast (30 hours. mean division time) as the uninfected controls or the cells infected with RAV-1 virus. At 36 degrees C the SR-RSV infected cells stop growing after the same delay as the controls, thus having undergone twice as many doublings, (38 to 40 instead 18 to 20). However, at 41 degrees C, SR-RSV infected cells stop 5 to 6 divisions earlier. This cannot be reversed by shifting the cultures to 36 degrees C. Rising to 41 degrees C those cultures which grew previously at 36 degrees C reduce the number of residual divisions. No such effect is observed with NYts68 or PA101 viruses which display a reduced level of pp60v-src activity at 41 degrees C.

Animals↗

PubMed1980