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Biomedical subjects

M Rocchi

Publications and source records attributed to M Rocchi.

At least 163 records · Page 9Linked to original sources
PubMed1994

Interspecies somatic T cell hybrids as biological tools for studying gene expression during T cell development.

Interspecies somatic cell hybrids were generated by fusing the mouse thymic lymphoma cell line, BW5147, with normal human T lymphocytes at different stages of differentiation. Thymocytes, activated peripheral T lymphocytes, or an activated T cell clone were used as human partners, respectively, in three independent fusions. Phenotype and genetic analysis demonstrated that these hybrids preferentially segregated human chromosomes while retaining a complete mouse genetic complement, irrespective of the human partner used for fusion. A large number of T cell differentiation antigens constitutively expressed throughout the T lymphocyte development remained constitutively expressed in the hybrids, irrespective of the maturation stage of human partner used for fusion. In contrast, the expression of other antigens related to a specific stage of T cell development (CD2, CD8), or to an activated state of T lymphocytes (HLA-DR, CD25), was to observed in the hybrids, with no apparent correlation with the segregation of human chromosomes other than, of course, the encoding chromosome. From these results we suggest that the developmental stage of the fusion partners strongly influences the pattern of expression by activating or silencing genes programmed to be expressed in distinct phases of T cell ontogeny.

Animals↗
PubMed1994

Seroepidemiological and clinical survey of feline immunodeficiency virus infection in northern Italy.

Four hundred and thirty-nine feline serum samples from cats with different living conditions in the north of Italy were tested for antibodies to feline immunodeficiency virus (FIV) and for antigen of Feline Leukemia Virus by enzyme-linked immunosorbent assay. A Western blot technique was also used on the positive sera in order to confirm the presence of specific antibodies to FIV. The Western blot enabled the detection of a false positive serum. The prevalence of FIV infection in this population was 12.5% and among the seropositive cats a greater proportion was male (74.5%) than female (25.5%). A correlation between the clinical status and the evolution of the pathology is described together with a score based on the severity of the stomatitis in infected cats. The Western blot patterns of positive samples were then compared with the stage of the pathology. Statistical analysis on the distribution of FIV in stray cats, cats with garden and courtyard access and strictly house-confined cats showed a highly significant risk of the infection in the first group.

Animals↗
PubMed1993

Cloning of human and rat cDNAs encoding the mitochondrial single-stranded DNA-binding protein (SSB).

We have retro-transcribed and amplified by PCR the full-length cDNAs specifying the rat and human precursors of the single-stranded mitochondrial DNA (mtDNA)-binding protein (mtSSB). Each deduced sequence is composed of a 16-amino-acid (aa) N-terminal basic pre-sequence and a mature protein (132 aa in humans and 135 aa in the rat). The mature proteins are highly conserved among themselves and with the mtSSB from Xenopus laevis (Xl). Moreover, three regions of the protein are similar to corresponding domains of the SSB of Escherichia coli and to the E. coli F-sex factor SSB, indicating the existence of a broad class of DNA-binding proteins with structural and functional similarities both in prokaryotes and in prokaryote-derived organelles of higher organisms.

Amino Acid Sequence↗
PubMed1993

Molecular organization and chromosomal location of human GC-rich heterochromatic blocks.

From the sequencing of three genomic DNA fragments and PCR amplification products from total human DNA, we have derived the sequence of a 545-bp Sau3A fragment (68% GC), representative of a family of human DNA repeats. Since previous studies suggested its linkage with unrelated Sau3A repeats of 68 bp (54% GC) (beta-satellite sequences), this feature was further investigated by in situ hybridization experiments and by Southern blot analysis of a panel of DNAs from human-Chinese hamster somatic cell hybrids. Both DNA repeats are preferentially localized on the heterochromatic regions of acrocentric chromosomes, on the pericentromeric heterochromatin of chromosome 1, 3 and 9, and on the proximal euchromatic region of the chromosome Y q arm. On chromosome 9, both repeats are part of a 2.7-kb higher-order repeat unit. These results and the Southern blot analysis on partial digests of total DNA, suggest that the linkage between the two repetitive DNA sequences is a constant feature throughout the genome. Furthermore, Southern blot analysis of HpaII-digested and MspI-digested DNA from different human tissues and tumor cell lines indicates that the investigated heterochromatic blocks appear to be subjected to changes in their methylation pattern.

Base Sequence↗
PubMed1993

Complementary DNA sequence and chromosomal mapping of a human proteoglycan-binding cell-adhesion protein (dermatopontin).

We have noticed the presence of a protein with a M(r) of approx 22 kDa in proteoglycan preparations from human fibroblast cultures and speculated that it might be related to a 22-kDa protein from bovine skin (22K) with proteoglycan- and cell-binding properties. Using degenerated oligomers designed from the amino acid sequence of the bovine protein, we amplified and subcloned sequences from human fibroblast and fibrosarcoma cDNA. The three clones that were characterized contain an open reading frame (603 bp) coding for 201 amino acids comprising a secretory leader peptide of 18 amino acids and a secreted part of 183 amino acids with 96% identity to the bovine sequence, indicating that they code for the human homologue ("dermatopontin") of the bovine 22K protein. Expression of dermatopontin is not limited to connective tissue, as Northern blots show specific mRNAs in cultured fibroblasts, muscle, heart, pancreas, and lung. Two species of mRNA (1.0 and 2.2 kb) are present, indicating alternative polyadenylation or alternative splicing. The cDNA clones map to 1q12-q23 in a cell hybrid panel containing specific chromosomal deletions.

Alternative Splicing↗
PubMed1993

Cloning and comparative mapping of a human chromosome 4-specific alpha satellite DNA sequence.

We have isolated and characterized two human alphoid DNA clones: p4n1/4 and pZ4.1. Clone p4n1/4 identifies specifically the centromeric region of chromosome 4; pZ4.1 recognizes a subset of alphoid DNA shared by chromosomes 4 and 9. The specificity was determined using fluorescence in situ hybridization experiments on metaphase spreads and Southern blotting analysis of human-hamster somatic cell hybrids. The genomic organization of both subsets was also investigated. Comparative mapping on chimpanzee and gorilla chromosomes was performed. p4n1/4 hybridizes to chimpanzee chromosomes 11 and 13, homologs of human chromosomes 9 and 2q, respectively. On gorilla metaphase spreads, p4n1/4 hybridizes exclusively to the centromeric region of chromosome 19, partially homologous to human chromosome 17. No hybridization signal was detected on chromosome 3 of both chimpanzee and gorilla, in both species homolog of human chromosome 4. Identical comparative mapping results were obtained using pZ4.1 probe, although the latter recognizes an alphoid subset distinct from the one recognized by p4n1/4. The implications of these results in the evolution of centromeric regions of primate chromosomes are discussed.

Animals↗
PubMed1993

Detection of a neurofibromatosis type I (NF1) homologous sequence by PCR: implications for the diagnosis and screening of genetic diseases.

The neurofibromatosis type I (NF1) gene was extensively screened for mutations using single strand conformation polymorphism (SSCP) technology. During the analysis of the NF1 GAP-related domain, electrophoretically abnormal fragments were detected. Direct sequencing of these fragments allowed us to identify the presence of a NF1 highly homologous sequence (NF1HHS). A detailed analysis of a hybrid panel located this sequence on chromosome 15q24-->qter. An accurate search through several data banks demonstrated that this sequence is a new NF1 homologue. This report shows how it is possible to find homologous sequences at random, and subsequently to make wrong interpretations.

Animals↗
PubMed1993

An alphoid DNA sequence conserved in all human and great ape chromosomes: evidence for ancient centromeric sequences at human chromosomal regions 2q21 and 9q13.

Using vector-CENP-B box polymerase chain reaction (PCR) we isolated and cloned from a human chromosome 21-specific plasmid library, a 1 kb DNA sequence, named p alpha H21. In in situ hybridization experiments, p alpha H21 hybridized, under high stringency conditions, to the centromeric region of all the human, chimpanzee, gorilla and orangutan chromosomes. On human chromosomes p alpha H21 also identified non-centromeric sequences at 2q21 (locus D2F33S1) and 9q13 (locus D9F33S2). The possible derivation of these sequences from ancestral centromeres is discussed. Sequence analysis confirmed the alphoid nature of the whole p alpha H21 insert.

Animals↗
PubMed1993

A novel X-linked member of the human zinc finger protein gene family: isolation, mapping, and expression.

We report the partial characterization of a novel putative zinc finger gene of the Krüppel-type (ZNF81), isolated from an X Chromosome (Chr) specific library. The pattern of segregation in human-hamster somatic cell hybrids of sequences homologous to the ZNF81 finger domain has established that it resides within the Xp22.1-Xp11 region. ZNF81 represents yet another example, together with ZFX, ZNF41, and ZNF21, of members of the zinc finger gene family residing within the short arm of the human X Chr. Sequence analysis showed that ZNF81 may encode a polypeptide(s) containing tandem arrays of 12 canonical C2H2 zinc fingers of the Krüppel-type at the C-terminus. Northern analysis indicated that probes from the ZNF81 finger domain hybridize to polyadenylated transcripts present in several cell lines, a result that supports the hypothesis that it is an expressed, functional member of this multigene family.

Amino Acid Sequence↗
PubMed1993

Lamin A/C gene and a related sequence map to human chromosomes 1q12.1-q23 and 10.

Lamins A and C are products of alternate splicing of one transcript from a single gene. We have isolated a partial cDNA, cE1-2, whose 800-bp sequence is 99% identical to the 3' untranslated region of the lamin A/C gene. We report here the mapping of this gene and a closely related sequence to human chromosomes 1 and 10, more specifically, to 1q12.1-q23. The localization of cE1-2 hybridizing sequences to two different chromosomes suggests that one of these loci represents an as yet unknown member of the lamin gene family, either a pseudogene or an expressed gene.

Animals↗
PubMed1993

A high resolution deletion map of human chromosome Xp22.

We have developed a 32-interval deletion panel for human chromosome Xp22 spanning about 30 megabases of genomic DNA. DNA samples from 50 patients with chromosomal rearrangements involving Xp22 were tested with 60 markers using a polymerase chain reaction strategy. The ensuing deletion map allowed us to confirm and refine the order of previously isolated and newly developed markers. Our mapping panel will provide the framework for mapping new sequences, for orienting chromosome walks in the region and for projects aimed at isolating genes responsible for diseases mapping to Xp22.

Base Sequence↗
PubMed1993

Mapping of two genes encoding isoforms of the actin binding protein ABP-280, a dystrophin like protein, to Xq28 and to chromosome 7.

ABP-280 is a ubiquitous actin binding protein present in the cytoskeleton of many different cell types. ABP-280 was mapped to distal Xq28, 50-60 kb downstream of the Green Colour Pigment (GCP) genes. To establish if ABP-280 may be a candidate for one of the muscle disease localized by linkage analysis to distal Xq28 we looked for alternative forms of ABP-280 mRNA. Several different ABP-280 mRNAs were indeed identified: two are X-linked and are produced by alternative splicing of a small exon of 24 nucleotides. At least one additional gene encoding a RNA more than 70% identical to ABP-280 in the 1700 bp sequenced has also been found. It was mapped to chromosome 7. While both forms of the X-linked ABP-280 are ubiquitous, the gene on chromosome 7 is highly expressed only in skeletal muscle and heart. The two genes were therefore excellent candidates for the X-linked and for the autosomal dominant form of the Emery-Dreifuss Muscular Dystrophy (EDMD) both of which have been described. So far, however we were unable to demonstrate mutations in the coding region or affecting the alternative splicing of the X-linked form of ABP-280, in several patients studied, and we think that it is quite unlikely that this is the gene responsible for EDMD.

Amino Acid Sequence↗
PubMed1993

Use of polymerase chain reaction to diagnose bovine leukemia virus infection in calves at birth.

A specific polymerase chain reaction (PCR) assay was devised, allowing detection of 1 bovine leukemia virus (BLV)-infected cell in 10(4) bovine lymphocytes. The efficacy of field application of the developed method was verified by evaluating the rate of viral transmission to calves from infected cows, whether they have persistent lymphocytosis. With this objective, 43 calves were simultaneously tested at birth and at 6 months of age for viral antibodies in serum and for proviral DNA in lymphocytes. At birth, 36 calves were BLV-negative and 3 were BLV-positive by results of serologic and DNA-based assays. Conversely, results for 4 calves had lack of correlation between the diagnostic methods. In particular, 2 calves were DNA-positive and antibody-negative for BLV and 2 other calves had the opposite test results. At 6 months of age, when the immunologic pattern more closely reflects the status of calves' immune response, independent of maternal antibodies, all calves DNA-negative for BLV at birth (n = 38), were consistently PCR- and antibody-negative for BLV. On the contrary, the cattle DNA-positive for BLV at birth (n = 5), whether seropositive or not, were PCR- and antibody-positive for BLV, at the time of the second screening. Thus, these results indicate reliability of the PCR to diagnose perinatal BLV infection. Furthermore, the observation that all calves found to be infected at birth were born to BLV-positive cows with persistent lymphocytosis, indicates that the persistent lymphocytosis status of the cow may represent a factor associated with BLV infection in utero.

Animals↗
PubMed1992

Characterization by yeast artificial chromosome cloning of the linked apolipoprotein(a) and plasminogen genes and identification of the apolipoprotein(a) 5' flanking region.

The apoprotein(a) [apo(a)] gene encodes a protein component of the circulating lipoprotein(a) [Lp(a)]. The apo(a) gene is highly homologous to the plasminogen gene. It encodes one of the most polymorphic human proteins, due to variability in the number of repetitions of structures called kringles. In addition, Lp(a) levels vary among individuals by more than two orders of magnitude, the high levels being highly correlated with predisposition to early atherosclerotic disease. To better understand the genetics and function of the apo(a) gene, we have cloned in yeast artificial chromosome vectors DNA fragments comprising the linked apo(a) and plasminogen genes and other members of the plasminogen family. By a combination of pulsed-field gel electrophoresis and genome walking experiments, we have identified the 5' portion and flanking regions of the apo(a) gene.

Base Sequence↗
PubMed1992

Interleukin-1-inducible genes in endothelial cells. Cloning of a new gene related to C-reactive protein and serum amyloid P component.

Differential screening of a cDNA library constructed from human umbilical vein endothelial cells exposed for 1 h to interleukin-1 beta (IL-1 beta) has led to the identification of a novel gene (PTX3) related to pentaxins (C-reactive protein and serum amyloid P component in man), a subclass of acute phase proteins. Sequencing of the full-length cDNA clone and RNase mapping revealed that the PTX3 transcript is 1861 base pairs long and has a unique transcription start site. The predicted protein sequence of 381 amino acids is highly similar to pentaxins in its COOH-terminal half where it also contains a typical 8-amino acid "pentaxin signature" sequence. The NH2-terminal half of PTX3 shows no similarity to any known protein sequence and initiates with a putative signal peptide indicating that PTX3 is secreted. The genome of PTX3 is organized into three exons. Interestingly, the region of homology between PTX3 and pentaxins corresponds to the third PTX3 exon. The PTX3 gene has been localized on human chromosome 3 band q25 by Southern blots of somatic cell hybrids and by in situ hybridization. The PTX3 mRNA is induced in endothelial, hepatic, and fibroblastic cells by IL-1 beta and tumor necrosis factor alpha but not by IL-6 and interferon-gamma. PTX3 may represent a novel marker of inflammatory reactions, particularly those involving the vessel wall.

Amino Acid Sequence↗
PubMed1992

The NM23 gene maps to human chromosome band 17q22 and shows a restriction fragment length polymorphism with BglII.

The NM23-Hl gene is a putative tumor suppressor gene that may be important in the metastasic process. Recent genetic and immunological data indicate that the NM23-Hl gene encodes a protein with nucleoside diphosphate (NDP) kinase activity. The mapping of NM23-Hl by panels of rodent-human somatic cell hybrids and in situ hybridization showed that the gene is located in human chromosome band 17q22. A two-allele polymorphism with BglII was demonstrated.

Alleles↗