PubMed Health⌕ Search

Biomedical subjects

M Wernick

Publications and source records attributed to M Wernick.

11 recordsLinked to original sources
PubMed2001

Evidence for alternate splicing within the mRNA transcript encoding the DNA damage response kinase ATR.

Proper cellular response to genotoxic insult often requires the activity of one or more members of a family of high-molecular weight protein kinases referred to as phosphatidylinositol-3 kinase (PIK)-like proteins. While catalytic activity is an indispensable part of PIK-like protein function, little is currently known about factors that control their activity and/or functions. This deficiency stems, in large part, from our lack of knowledge concerning functionally significant subdomains within the large non-catalytic domain of these proteins. We have determined that the transcript encoding the PIK-like protein ATR undergoes alternate splicing within the region of the mRNA encoding its non-catalytic domain. This conclusion is based on the sequencing of a human expressed sequence tag clone encoding a portion of the ATR cDNA, and is supported by the results of reverse transcriptase-polymerase chain reaction (RT-PCR) assays conducted on total and polyA+ RNA, as well as sequencing of cloned RT-PCR products. Cloning and sequencing of a segment of human genomic DNA indicated that this event arises from splicing of a single 192 bp exon within the ATR gene. Analysis of several human tissues indicated that alternate ATR transcripts are differentially expressed, suggesting that this region of the ATR protein may be of functional importance.

Alternative Splicing↗
PubMed2001

DNA polymerase eta undergoes alternative splicing, protects against UV sensitivity and apoptosis, and suppresses Mre11-dependent recombination.

Polymerase eta (pol eta) is a low-fidelity DNA polymerase that is the product of the gene, POLH, associated with the human XP variant disorder in which there is an extremely high level of solar-induced skin carcinogenesis. The complete human genomic sequence spans about 40 kb containing 10 coding exons and a cDNA of 2.14 kb; exon I is untranslated and is 6 kb upstream from the first coding exon. Using bacterial artificial chromosomes (BACs), the gene was mapped to human chromosome band 6p21 and mouse band 17D. The gene is expressed in most tissues, except for very low or undetectable levels in peripheral lymphocytes, fetal spleen, and adult muscle; exon II, however, is frequently spliced out in normal cells and in almost half the transcripts in the testis and fetal liver. Expression of POLH in a multicopy episomal vector proved nonviable, suggesting that overexpression is toxic. Expression from chromosomally integrated linear copies using either an EF1-alpha or CMV promoter was functional, resulting in cell lines with low or high levels of pol eta protein, respectively. Point mutations in the center of the gene and in a C-terminal cysteine and deletion of exon II resulted in inactivation, but addition of a terminal 3 amino acid C-terminal tag, or an N- or C-terminal green fluorescent protein, had no effect on function. A low level of expression of pol eta eliminated hMre11 recombination and partially restored UV survival, but did not prevent UV-induced apoptosis, which required higher levels of expression. Polymerase eta is therefore involved in S-phase checkpoint and signal transduction pathways that lead to arrest in S, apoptosis, and recombination. In normal cells, the predominant mechanism of replication of UV damage involves pol eta-dependent bypass, and Mre11-dependent recombination that acts is a secondary, backup mechanism when cells are severely depleted of pol eta.

Alternative Splicing↗
PubMed2001

Genome scanning with array CGH delineates regional alterations in mouse islet carcinomas.

Carcinomas that develop in the pancreatic islets of transgenic mice expressing the SV40 T-antigens (Tag) under transcriptional control of the rat insulin II promoter (RIP) progress through well-characterized stages that are similar to aspects of human tumor progression, including hyperplastic growth, increased angiogenesis and reduced apoptosis. The latter two stages have been associated with recurrent loss of heterozygosity (LOH) and reduced genome copy number on chromosomes 9 (LOH9) and 16 (LOH16), aberrations which we believe contribute to these phenotypes. Earlier analyses localized LOH9 to approximately 3 Mb and LOH16 to approximately 30 Mb (both syntenic with human 3q21-q25) but were limited by low throughput and a lack of informative polymorphic markers. Here we show that comparative genomic hybridization to DNA microarrays (array CGH) overcomes these limitations by allowing efficient, genome-wide analyses of relative genome copy number. The CGH arrays used in these experiments carried BACs distributed at 2-20-MB intervals across the mouse genome and at higher density in regions of interest. Using array CGH, we further narrowed the loci for LOH9 and LOH16 and defined new or previously unappreciated recurrent regions of copy-number decrease on chromosomes 6, 8 and 14 (syntenic with human chromosomes 12p11-p13, 16q24.3 and 13q11-q32, respectively) and regions of copy-number increase on chromosomes 2 and 4 (syntenic to human chromosomes 20q13.2 and 1p32-p36, respectively). Our analyses of human genome sequences syntenic to these regions suggest that CYP24, PFDN4, STMN1, CDKN1B, PPP2R3 and FSTL1 are candidate oncogenes or tumor-suppressor genes. We also show that irradiation and genetic background influence the spectrum of aberrations present in these tumors.

Animals↗
PubMed2001

Comprehensive genome sequence analysis of a breast cancer amplicon.

Gene amplification occurs in most solid tumors and is associated with poor prognosis. Amplification of 20q13.2 is common to several tumor types including breast cancer. The 1 Mb of sequence spanning the 20q13.2 breast cancer amplicon is one of the most exhaustively studied segments of the human genome. These studies have included amplicon mapping by comparative genomic hybridization (CGH), fluorescent in-situ hybridization (FISH), array-CGH, quantitative microsatellite analysis (QUMA), and functional genomic studies. Together these studies revealed a complex amplicon structure suggesting the presence of at least two driver genes in some tumors. One of these, ZNF217, is capable of immortalizing human mammary epithelial cells (HMEC) when overexpressed. In addition, we now report the sequencing of this region in human and mouse, and on quantitative expression studies in tumors. Amplicon localization now is straightforward and the availability of human and mouse genomic sequence facilitates their functional analysis. However, comprehensive annotation of megabase-scale regions requires integration of vast amounts of information. We present a system for integrative analysis and demonstrate its utility on 1.2 Mb of sequence spanning the 20q13.2 breast cancer amplicon and 865 kb of syntenic murine sequence. We integrate tumor genome copy number measurements with exhaustive genome landscape mapping, showing that amplicon boundaries are associated with maxima in repetitive element density and a region of evolutionary instability. This integration of comprehensive sequence annotation, quantitative expression analysis, and tumor amplicon boundaries provide evidence for an additional driver gene prefoldin 4 (PFDN4), coregulated genes, conserved noncoding regions, and associate repetitive elements with regions of genomic instability at this locus.

Animals↗
PubMed1999

Preliminary assessment of extrastriatal dopamine D-2 receptor binding in the rodent and nonhuman primate brains using the high affinity radioligand, 18F-fallypride.

We have identified the value of 18F-fallypride [(S)-N-[(1-allyl-2-pyrrolidinyl)methyl]-5-(3-[18F]fluoropropyl)-2, 3-dimethoxybenzamide], as a dopamine D-2 receptor radiotracer for the study of striatal and extrastriatal receptors. Fallypride exhibits high affinities for D-2 and D-3 subtypes and low affinity for D-4 (3H-spiperone IC50s: D-2 = 0.05 nM [rat striata], D-3 = 0.30 nM [SF9 cell lines, rat recombinant], and D-4 = 240 nM [CHO cell lines, human recombinant]). Biodistribution in the rat brain showed localization of 18F-fallypride in striata and extrastriatal regions such as the frontal cortex, parietal cortex, amygdala, hippocampus, thalamus, and hypothalamus. In vitro autoradiographic studies in sagittal slices of the rat brain showed localization of 18F-fallypride in striatal and several extrastriatal regions, including the medulla. Positron emission tomography (PET) experiments with 18F-fallypride in male rhesus monkeys were carried out in a PET VI scanner. In several PET experiments, apart from the specific binding seen in the striatum, specific binding of 18F-fallypride was also identified in extracellular regions (in a lower brain slice, possibly the thalamus). Specific binding in the extrastriata was, however, significantly lower compared with that observed in the striata of the monkeys (extrastriata/cerebellum = 2, striata/cerebellum = 10). Postmortem analysis of the monkey brain revealed significant 18F-fallypride binding in the striata, whereas binding was also observed in extrastriatal regions such as the thalamus, cortical areas, and brain stem.

Animals↗
PubMed1999

Exercise--it's never too late: the strong-for-life program.

OBJECTIVES: This investigation determined whether an in-home resistance training program achieved health benefits in older adults with disabilities. METHODS: A randomized controlled trial compared the effects of assigning 215 older persons to either a home-based resistance exercise training group or a waiting list control group. Assessments were conducted at baseline and at 3 and 6 months following randomization. The program consisted of videotaped exercise routines performed with elastic bands of varying thickness. RESULTS: High rates of exercise adherence were achieved, with 89% of the recommended exercise sessions performed over 6 months. Relative to controls, subjects who participated in the program achieved statistically significant lower extremity strength improvements of 6% to 12%, a 20% improvement in tandem gait, and a 15% to 18% reduction in physical and overall disability at the 6-month follow-up. No adverse health effects were encountered. CONCLUSIONS: These findings provide important evidence that home-based resistance exercise programs designed for older persons with disabilities hold promise as an effective public health strategy.

Aged↗
PubMed1998

The human homologue for the Caenorhabditis elegans cul-4 gene is amplified and overexpressed in primary breast cancers.

Amplification is a key mechanism whereby a cancer cell increases the message level of genes that confer a selective advantage when they are overexpressed. In breast cancer, there are many chromosome regions present in multiple copies relative to overall DNA copy number (amplicons), and their target genes are unknown. Using differential display, we have cloned and sequenced the full coding region of a candidate amplicon target gene located on chromosome 13. This candidate is the human homologue of the Caenorhabditis elegans cul-4 gene, cul-4A, a member of the novel cullin gene family, which is involved in cell cycle control of C. elegans. cul-4A was amplified and overexpressed in 3 of 14 breast cancer cell lines analyzed, and it was overexpressed in 8 additional cell lines in which it was not amplified. The latter observation, indicating that its overexpression can occur by mechanisms other than gene amplification, suggests that cul-4A plays a key role in carcinogenesis. Moreover, cul-4A was found to be amplified in 17 of 105 (16%) cases of untreated primary breast cancers, and 14 of 30 cases analyzed (47%) were shown by RNA in situ hybridization to overexpress cul-4A. These results suggest that up-regulation of cul-4A may play an important role in tumor progression.

Amino Acid Sequence↗
PubMedDate not supplied

"Learned helplessness" and the patient with progressive supranuclear palsy.

Progressive supra-nuclear palsy is a neurologically debilitating disease of unknown cause and with no effective treatment. Learned helplessness, a major cause of depression in the institutionalized elderly, is a feeling of worthlessness and powerlessness resulting in a decreased incentive to try. This article provides insight into these two problems and specific nursing interventions to enhance self care.

Aged↗