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I Horak

Publications and source records attributed to I Horak.

15 recordsLinked to original sources
PubMed1992

Expression of human foamy virus is differentially regulated during development in transgenic mice.

The human foamy virus (HFV) is a recently characterized member of the spumavirus family. Although no diseases have been unequivocally associated with HFV infection, expression of HFV regulatory genes in transgenic mice induces a characteristic acute neurodegenerative disease and a myopathy. To better characterize the sequence of events leading to disease, and to gain a better understanding of the underlying pathogenetic mechanisms, we have analyzed in detail the transgene expression pattern during development. Transcription of a construct containing all regulatory elements and ancillary genes of HFV was analyzed by in situ hybridization and was shown to occur in two distinct phases. At midgestation, low but widespread expression was first detected in cells of extraembryonic tissues. Later, various tissues originating from embryonic mesoderm, neuroectoderm, and neural crest transcribed the transgene at moderate levels. However, expression decreased dramatically during late gestation and was suppressed shortly after birth. After a latency period of up to 5 weeks, transcription of the transgene resumed in single cells distributed irregularly in the central nervous system and in the skeletal muscle. By the age of 8 weeks, an increasing number of cells displayed much higher expression levels than in embryonic life and eventually underwent severe degenerative changes. These findings demonstrate that HFV transgene expression is differentially regulated in development and that HFV cytotoxicity may be dose-dependent. Such biphasic pattern of expression differs from that of murine retroviruses and may be explained by the specificity of HFV regulatory elements in combination with cellular factors. Future studies of this model system should, therefore, provide novel insights in the mechanisms controlling retroviral latency.

Animals↗
PubMed1992

Human foamy virus: an underestimated neuropathogen?

Human foamy virus (HFV) is a recently characterized retrovirus which was originally isolated from patients with various neoplastic and degenerative diseases. However, until today it has not been possible to identify HFV as the causative agent of any disease and little is known about its prevalence in human populations. Like HTLV and HIV, HFV encodes the three structural retroviral genes, gag, pol and env, and an additional region containing three open reading frames, bel-1 to bel-3. Bel-1 activates transcription of the long terminal repeat of HFV and HIV. In order to study the consequences of expressing HFV regulatory genes and to investigate a possible pathogenic potential of HFV, we have introduced parts of the HFV genome into the germ line of mice. Our studies with transgenic mice demonstrate that HFV transgenes encompassing the bel region are transiently transcribed between midgestation and birth at moderate levels in various tissues. Expression is then suppressed, but resumes after a latency of several weeks in a restricted range of tissues and leads to extensive accumulation of HFV transcripts in single cells. This is associated with a progressive degenerative disease of the central nervous system and of striated muscle. These findings provide the first evidence of a disease induced by HFV and suggest that HFV might also act as a human pathogen in neurological diseases. Moreover, the transgenic mouse model will be useful for studying the molecular basis of HFV-induced neurotoxicity, the role of individual disease-associated HFV genes and the regulation of retroviral latency.

Animals↗
PubMed1991

Development and function of T cells in mice rendered interleukin-2 deficient by gene targeting.

Interleukin-2 (IL-2) is a lymphocytotropic hormone which is thought to have a key role in the immune response of mammalian cells. It is produced by a subpopulation of activated T-lymphocytes and acts in vitro as the principal auto- and paracrine T-cell growth factor (for reviews see refs 1-3). IL-2 is, however, not the sole T-cell growth factor, nor does it act exclusively on T cells, also promoting growth of NK cells and differentiation of B cells. A role for IL-2 in T-cell development has been postulated but remains controversial. Here we test the requirement for IL-2 in vivo using IL-2-deficient mice generated by targeted recombination. We find that mice homozygous for the IL-2 gene mutation are normal with regard to thymocyte and peripheral T-cell subset composition, but that a dysregulation of the immune system is manifested by reduced polyclonal in vitro T-cell responses and by dramatic changes in the isotype levels of serum immunoglobulins.

Animals↗
PubMed1991

Progressive encephalopathy and myopathy in transgenic mice expressing human foamy virus genes.

Transgenic mice carrying the bel region of human foamy retrovirus (HFV) under transcriptional control of its own long terminal repeat expressed the transgene in their central nervous systems and in smooth and striated muscle tissues. The animals developed a progressive degenerative disease of the central nervous system and of the striated muscle. Because expression of the transgene was closely correlated with the appearance of structural damage and inflammatory reactions were scanty, the disease is likely to be caused directly by the HFV proteins. These unexpected findings call for a reevaluation of the pathogenic potential of HFV in humans.

Animals↗
PubMed1989

A recombination hotspot in the LTR of a mouse retrotransposon identified in an in vitro system.

The recombinational frequency between two long terminal repeat elements (LTR-IS) of a mouse retrotransposon was about 13 times higher, compared with that of two control DNA sequences in extracts from mouse testes, but not in extracts from ascites cells. Deletion of a 37 bp region from the LTR-IS element strongly suppresses its recombinational activity. This 37 bp region encompasses an area of potentially single-stranded DNA and interacts with at least two nuclear proteins. One of them binds sequence-specifically to single-stranded DNA and is present in both types of extracts. Another protein(s) binds to dsDNA at the motif TGGAAATCCCC and is absent in extracts from testes. Our results suggest that a cis-acting DNA sequence within the 504 bp LTR-IS element is responsible for its high recombinational activity in vitro, and they further support the previous suggestion that the LTR-IS elements are meiotic recombinational hotspots in vivo.

Animals↗
PubMed1987

Isolation of novel human retrovirus-related sequences by hybridization to synthetic oligonucleotides complementary to the tRNA(Pro) primer-binding site.

Synthetic oligonucleotides complementary to putative retroviral primer-binding sites were used as hybridization probes to detect novel retroviruslike sequences. An 8.1-kilobase element with structural features of a retroviral provirus was isolated from a human genomic library by this approach. Nucleotide sequence analysis of its 600-base-pair long terminal repeats revealed characteristic motifs known as regulatory signals for RNA polymerase II transcription: CCAAT, TATA, and ATTAAA. In addition, a putative pol gene displays apparent homologies to conserved regions of retroviral reverse transcriptase. The 5' long terminal repeat is flanked at its 3' end by a putative primer-binding site for reverse transcription with homology to tRNA(Pro). This element is therefore termed HuRRS-P (human retrovirus-related sequence-proline). There are 20 to 40 copies of HuRRS-P homologous sequences in DNAs of human and simian origin.

Animals↗
PubMed1978

Identification and characterization of ribosomal proteins phosphorylated in vaccinia-virus-infected HeLa cells.

Two-dimensional analysis of 32P-labelled ribosomal proteins revealed three proteins which are phosphorylated in vaccinia-virus-infected HeLa cells. All three proteins belong to the 40-S ribosomal subunits and were identified as S2, S6 and S16. The ribosomal protein S6 is phosphorylated also in uninfected HeLa cells. Phosphoserine was detected in all three proteins, phosphothreonine only in the protein S2. Phosphorylation of these ribosomal proteins in infected cells is dependent on the multiplicity of the viral infection and increases during the first six hours of infection. All three proteins are also phosphorylated in virus-infected cells treated with cycloheximide and in cells infected with ultraviolet-irradiated virus. This suggests that the phosphorylation reaction involves a vaccinia virion-associated protein kinase.

Amino Acids↗
PubMed1978

Ribosomal proteins of HeLa cells.

Ribosomal proteins from HeLa cells were analyzed by two-dimensional polyacrylamide gel electrophoresis (Kaltschmidt-Wittmann) and dodecylsulfate polyacrylamide gel electrophoresis (Laemmli). 35 proteins are associated with the small ribosomal subunit and 47 proteins with the large ribosomal subunit. The HeLa ribosomal proteins S6, S32, L40b,c, L41 and L42 are phosphorylated in vivo and in vitro. Minor differences between HeLa and rat liver ribosomal proteins were revealed by their direct coelectrophoresis.

Animals↗
PubMed1977

Acidic phosphoproteins of the 60-S ribosomal subunits from HeLa cells.

Two-dimensional analysis of the ribosomal proteins from 60-S subunits of HeLa cells revealed a triplet of acidic proteins, L40a, L40b and L40c, of identical molecular weight (13,700), which can be separated only on the basis of their charge differences. Two of the spots, L40b and L40c, become labeled after incubation of the cells with inorganic [32P]phosphate. The electrophoretic behavior and molecular weights of these proteins support the notion that the proteins L40b and L40c, are phosphorylated forms of the protein L40a. The same proteins can be phosphorylated also in vitro by a HeLa protein kinase on 60-S subunits but not on 80-S ribosomes. The inaccessibility of L40 proteins to the phosphorylation in vitro on 80-S ribosomes suggests that they are located in the interface between the 40-S and 60-S subunits.

Electrophoresis, Polyacrylamide Gel↗