Multiple Pathway-Based Genetic Variations Associated with Tobacco Related Multiple Primary Neoplasms

BACKGROUND: In order to elucidate a combination of genetic alterations that drive tobacco carcinogenesis we have explored a unique model system and analytical method for an unbiased qualitative and quantitative assessment of gene-gene and gene-environment interactions. The objective of this case control study was to assess genetic predisposition in a biologically enriched clinical model system of tobacco related cancers (TRC), occurring as Multiple Primary Neoplasms (MPN). METHODS: Genotyping of 21 candidate Single Nucleotide Polymorphisms (SNP) from major metabolic pathways was performed in a cohort of 151 MPN cases and 210 cancer-free controls. Statistical analysis using logistic regression and Multifactor Dimensionality Reduction (MDR) analysis was performed for studying higher order interactions among various SNPs and tobacco habit. RESULTS: Increased risk association was observed for patients with at least one TRC in the upper aero digestive tract (UADT) for variations in SULT1A1 Arg²¹³His, mEH Tyr¹¹³His, hOGG1 Ser³²⁶Cys, XRCC1 Arg²⁸⁰His and BRCA2 Asn³⁷²His. Gene-environment interactions were assessed using MDR analysis. The overall best model by MDR was tobacco habit/p53(Arg/Arg)/XRCC1(Arg³⁹⁹His)/mEH(Tyr¹¹³His) that had highest Cross Validation Consistency (8.3) and test accuracy (0.69). This model also showed significant association using logistic regression analysis. CONCLUSION: This is the first Indian study on a multipathway based approach to study genetic susceptibility to cancer in tobacco associated MPN. This approach could assist in planning additional studies for comprehensive understanding of tobacco carcinogenesis

Alternative Splicing in the Differentiation of Human Embryonic Stem Cells into Cardiac Precursors

The role of alternative splicing in self-renewal, pluripotency and tissue lineage specification of human embryonic stem cells (hESCs) is largely unknown. To better define these regulatory cues, we modified the H9 hESC line to allow selection of pluripotent hESCs by neomycin resistance and cardiac progenitors by puromycin resistance. Exon-level microarray expression data from undifferentiated hESCs and cardiac and neural precursors were used to identify splice isoforms with cardiac-restricted or common cardiac/neural differentiation expression patterns. Splice events for these groups corresponded to the pathways of cytoskeletal remodeling, RNA splicing, muscle specification, and cell cycle checkpoint control as well as genes with serine/threonine kinase and helicase activity. Using a new program named AltAnalyze (http://www.AltAnalyze.org), we identified novel changes in protein domain and microRNA binding site architecture that were predicted to affect protein function and expression. These included an enrichment of splice isoforms that oppose cell-cycle arrest in hESCs and that promote calcium signaling and cardiac development in cardiac precursors. By combining genome-wide predictions of alternative splicing with new functional annotations, our data suggest potential mechanisms that may influence lineage commitment and hESC maintenance at the level of specific splice isoforms and microRNA regulation

Inhibition of protein ubiquitination by paraquat and 1-methyl-4-phenylpyridinium impairs ubiquitin-dependent protein degradation pathways

Intracytoplasmic inclusions of protein aggregates in dopaminergic cells (Lewy bodies) are the pathological hallmark of Parkinson’s disease (PD). Ubiquitin (Ub), alpha [α]-synuclein, p62/sequestosome 1 and oxidized proteins are major components of Lewy bodies. However, the mechanisms involved in the impairment of misfolded/oxidized protein degradation pathways in PD are still unclear. PD is linked to mitochondrial dysfunction and environmental pesticide exposure. In this work, we evaluated the effect of the pesticide paraquat (PQ) and the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP+) on Ub-dependent protein degradation pathways. No increase in the accumulation of Ub-bound proteins or aggregates was observed in dopaminergic cells (SK-N-SH) treated with PQ or MPP+, or in mice chronically exposed to PQ. PQ decreased Ub protein content, but not its mRNA transcription. Protein synthesis inhibition with cycloheximide depleted Ub levels and potentiated PQ–induced cell death. Inhibition of proteasomal activity by PQ was found to be a late event in cell death progression, and had no effect on either the toxicity of MPP+ or PQ, or the accumulation of oxidized sulfenylated, sulfonylated (DJ-1/PARK7 and peroxiredoxins) and carbonylated proteins induced by PQ. PQ- and MPP+-induced Ub protein depletion prompted the dimerization/inactivation of the Ub-binding protein p62 that regulates the clearance of ubiquitinated proteins by autophagic. We confirmed that PQ and MPP+ impaired autophagy flux, and that the blockage of autophagy by the overexpression of a dominant-negative form of the autophagy protein 5 (dnAtg5) stimulated their toxicity, but there was no additional effect upon inhibition of the proteasome. PQ induced an increase in the accumulation of α-synuclein in dopaminergic cells and membrane associated foci in yeast cells. Our results demonstrate that inhibition of protein ubiquitination by PQ and MPP+ is involved in the dysfunction of Ub-dependent protein degradation pathways

A multimodal cell census and atlas of the mammalian primary motor cortex

ABSTRACT We report the generation of a multimodal cell census and atlas of the mammalian primary motor cortex (MOp or M1) as the initial product of the BRAIN Initiative Cell Census Network (BICCN). This was achieved by coordinated large-scale analyses of single-cell transcriptomes, chromatin accessibility, DNA methylomes, spatially resolved single-cell transcriptomes, morphological and electrophysiological properties, and cellular resolution input-output mapping, integrated through cross-modal computational analysis. Together, our results advance the collective knowledge and understanding of brain cell type organization: First, our study reveals a unified molecular genetic landscape of cortical cell types that congruently integrates their transcriptome, open chromatin and DNA methylation maps. Second, cross-species analysis achieves a unified taxonomy of transcriptomic types and their hierarchical organization that are conserved from mouse to marmoset and human. Third, cross-modal analysis provides compelling evidence for the epigenomic, transcriptomic, and gene regulatory basis of neuronal phenotypes such as their physiological and anatomical properties, demonstrating the biological validity and genomic underpinning of neuron types and subtypes. Fourth, in situ single-cell transcriptomics provides a spatially-resolved cell type atlas of the motor cortex. Fifth, integrated transcriptomic, epigenomic and anatomical analyses reveal the correspondence between neural circuits and transcriptomic cell types. We further present an extensive genetic toolset for targeting and fate mapping glutamatergic projection neuron types toward linking their developmental trajectory to their circuit function. Together, our results establish a unified and mechanistic framework of neuronal cell type organization that integrates multi-layered molecular genetic and spatial information with multi-faceted phenotypic properties

Preclinical studies for gene therapy of head and neck cancers using the HSV-tk/GCV strategy

82-87The present study has used the suicide gene Herpes Simplex Virus–thymidine kinase (HSV-tk) and ganciclovir (GCV) strategy. A 2 kb sequence of HSV-tk was subcloned into the retroviral vector resulting in a recombinant vector, LTKSN, which was transfected into a packaging cell-line, PA317 and selected on G418. The highest expressing clone, PTK-16 was used for in vitro experiments. Intra-tumoural injections of PTK-16 followed by GCV treatment, in a HNSCC xenograft model in nude mice, showed a significant reduction in the viable tumour volume (p=0.009). These results will form the basis for future clinical trials in HNSCC

Regulation of Apoptosis and Caspase-8 Expression in Neuroblastoma Cells by Isoforms of the IG20

Pavilion of Ten Thousand Autumns (Qian Qiu Ting) in the west garden; The Forbidden City was the Chinese imperial palace from the Ming Dynasty to the end of the Qing Dynasty. To the west of the Inner Palace is the residential quarter of the Qing emperors, with numerous smaller buildings and gardens. To the east stood the residence of the imperial concubines and various ritual halls, as well as theatres, temples and gardens. Source: Wikipedia; http://en.wikipedia.org/wiki/Main_Page (accessed 5/11/2011