Biomedical subjects

Sachin Katyal

Publications and source records attributed to Sachin Katyal.

5 recordsLinked to original sources

PubMed2006

The neurodegenerative disease protein aprataxin resolves abortive DNA ligation intermediates.

Ataxia oculomotor apraxia-1 (AOA1) is a neurological disorder caused by mutations in the gene (APTX) encoding aprataxin. Aprataxin is a member of the histidine triad (HIT) family of nucleotide hydrolases and transferases, and inactivating mutations are largely confined to this HIT domain. Aprataxin associates with the DNA repair proteins XRCC1 and XRCC4, which are partners of DNA ligase III and ligase IV, respectively, suggestive of a role in DNA repair. Consistent with this, APTX-defective cell lines are sensitive to agents that cause single-strand breaks and exhibit an increased incidence of induced chromosomal aberrations. It is not, however, known whether aprataxin has a direct or indirect role in DNA repair, or what the physiological substrate of aprataxin might be. Here we show, using purified aprataxin protein and extracts derived from either APTX-defective chicken DT40 cells or Aptx-/- mouse primary neural cells, that aprataxin resolves abortive DNA ligation intermediates. Specifically, aprataxin catalyses the nucleophilic release of adenylate groups covalently linked to 5'-phosphate termini at single-strand nicks and gaps, resulting in the production of 5'-phosphate termini that can be efficiently rejoined. These data indicate that neurological disorders associated with APTX mutations may be caused by the gradual accumulation of unrepaired DNA strand breaks resulting from abortive DNA ligation events.

Adenosine Monophosphate↗

PubMed2005

Dynamic nature of cleavage bodies and their spatial relationship to DDX1 bodies, Cajal bodies, and gems.

DDX1 bodies, cleavage bodies, Cajal bodies (CBs), and gems are nuclear suborganelles that contain factors involved in RNA transcription and/or processing. Although all four nuclear bodies can exist as distinct entities, they often colocalize or overlap with each other. To better understand the relationship between these four nuclear bodies, we examined their spatial distribution as a function of the cell cycle. Here, we report that whereas DDX1 bodies, CBs and gems are present throughout interphase, CPSF-100-containing cleavage bodies are predominantly found during S and G2 phases, whereas CstF-64-containing cleavage bodies are primarily observed during S phase. All four nuclear bodies associate with each other during S phase, with cleavage bodies colocalizing with DDX1 bodies, and cleavage bodies/DDX1 bodies residing adjacent to gems and CBs. Although inhibitors of RNA transcription had no effect on DDX1 bodies or cleavage bodies, inhibitors of DNA replication resulted in loss of CstF-64-containing cleavage bodies. A striking effect on nuclear structures was observed with latrunculin B, an inhibitor of actin polymerization, resulting in the formation of needlelike nuclear spicules made up of CstF-64, CPSF-100, RNA, and RNA polymerase II. Our results suggest that cleavage body components are highly dynamic in nature.

Actins↗

PubMed2004

Alternative splicing modulates Disabled-1 (Dab1) function in the developing chick retina.

The Reelin-Disabled 1 (Dab1)-signaling pathway plays a critical role in neuronal cell positioning in the brain. We have isolated two alternatively spliced variants of Dab1 from chick retina, an early form (chDab1-E) expressed in undifferentiated cells and a late form (chDab1-L) expressed in amacrine and ganglion cells. A key difference between the two forms is the exclusion in chDab1-E of two Src-related tyrosine kinase recognition sites implicated in Reelin-mediated Dab1 tyrosine phosphorylation. Retinal cultures transfected with a chDab1-L expression construct undergo a dramatic change in morphology, accompanied by the formation of numerous thin elongated processes, increased tyrosine phosphorylation, activation of Src family kinase(s) and increased levels of the axonal outgrowth protein growth-associated protein-43. In contrast, chDab1-E transfectants retain an undifferentiated morphology. Mutational analysis implicates a specific tyrosine (tyr-198) in the morphological and biochemical alterations associated with chDab1-L expression. We propose that alternative splicing of chDab1 represents an effective and flexible way of regulating the Reelin-Dab1-signaling pathway in a mixed cell population, by ensuring that secreted Reelin activates the signaling cascade only in target neuronal cells.

Alternative Splicing↗

PubMed2003

Isolation of a novel cDNA enriched in the undifferentiated chick retina and lens.

By using a differential screening strategy, we have identified a previously uncharacterised gene that is preferentially expressed in chick retinal precursor cells as well as in the anterior epithelial cells of the lens at early stages of development. The EURL transcript has an open reading frame of 293 amino acids and a 3' untranslated region of >850 nucleotides. The only recognizable feature in the EURL (early undifferentiated retina and lens) amino acid sequence is a putative coiled-coil domain located at the C-terminus of the protein. The human EURL orthologue maps to chromosome 21q21. Northern blot analysis, in situ hybridization of tissue sections, and whole-mount hybridization indicate elevated levels of EURL mRNA in the embryonic dorsal retina from stage 14 (day 2) to stage 18 (day 3). At day 3.5, when >90% of the retinal cells are in the proliferative stage, EURL transcripts are found primarily in the peripheral dorsal retina, i.e., the most undifferentiated part of the dorsal retina. EURL transcripts are also detected in the lens at stage 18 and remain abundant in the proliferating epithelial cells of the lens until at least day 11. The distribution pattern of EURL in the developing retina and lens suggest a role before the events leading to cell determination and differentiation.

3' Untranslated Regions↗

PubMed2002

Cloning and expression analysis of the chicken DEAD box gene DDX1.

DEAD box proteins are putative RNA unwinding proteins found in organisms ranging from mammals to bacteria. While some DEAD box genes expressed in higher eukaryotes are ubiquitous, others have distribution profiles that suggest a cell-, tissue-, or developmental-specific role. The DEAD box gene, DDX1, was identified by differential screening of a subtracted retinoblastoma cDNA library. A limited survey of human fetal tissues indicated that DDX1 mRNA has a widespread distribution but is not uniformly expressed in all tissues. To further document the spatial and temporal distribution of DDX1 during embryonic development, we cloned the chicken DDX1 cDNA. The predicted amino acid sequence of chicken DDX1 was 93% identical to that of human DDX1. All DEAD box motifs, as well as a SPRY domain, were present in chicken DDX1. Northern and Western blot analyses showed highest levels of DDX1 at early stages of development. Tissue maturation was generally accompanied by a decrease in expression, although DDX1 levels remained elevated in late embryonic retina and brain. In situ hybridization of retinal tissue sections revealed widespread distribution of DDX1 mRNA at early developmental stages with preferential expression in amacrine and ganglion cells of the differentiated tissue. Preferential expression of DDX1 was also observed in specific areas of the brain in older embryos, such as the external granule layer of the cerebellum. These results suggest a specific role for DDX1 in subsets of differentiated cells as well as a more general role in undifferentiated cells.

Amino Acid Sequence↗