Project description:PTEN Harmatoma Tumor Syndrome is a collection of clinical diagnosis with patients harbouring germline mutation in the PTEN gene. Most affected individuals have extreme macrocephaly and autism spectrum disorders were diagnosed in ~10% of the patients. However, the underpinning molecular mechanisms are not fully defined. The aim of this study is to compare the transcriptomic profiles of somatosensory cortices from Pten+/- and wild-type littermates (C57Bl6/J) of female and male at P30 and P42, primary neural cells, neuroprogenitor cells (NPCs), cortical astrocytes (PCA) and cortical neurons (PCN) from C57Bl6/J mice, with wild-type (WT as control for Pten+/- or CTRL as control for Nes-KO), heterozygous (Pten+/-)and complete knockout of Pten (Nes-KO). The study included performing RNA sequencing of these tissues and neural cells and integrating the data with human autism candidate genes (SFARI), and enrichment with GWAS genes for neurological diseases and traits. Methods: RNA sequencing of the somatosensory cortices and primary neural cells: Somatosensory cortices are cut from female and male mice at P30 and P42. NPCs are from E12.5 embryons. PCNs are from E15.5 embryos. PCAs are from P0 or P2 pups. Samples were lysed in TRIzol and RNA was extracted with chloroform/isopropanol and ethanol. Poly-A eniched RNA was used for library contruction. Results: We identified widespread signature genes in different samples, with enhanced immune response and oligodendrocyte development in somatosensory cortex, suppressed immediate early response genes in Pten+/- neurons, upregulated cilium components in astrocyte and downregulated inhibitory neurons in Pten knockout neural cells through RNA sequencing, which further enriched in GWAS genes of schizophrenia. Conclusion: Our study represents the first detailed comprehensive RNA-sequencing of somatosensory cortex and individual types of neural cells of transgenic mice with Pten heterozygous and homozygous knockout. The result suggested Pten effects on dysregulated immune response, immediately early response, cilium function and interneurons development which would direct further molecular studies on neuropathologenic mechanisms for PHTS.

Project description:Anaplasma phagocytophilum is an emerging zoonotic pathogen that causes human granulocytic anaplasmosis. These intracellular bacteria establish infection by affecting cell function in both the vertebrate host and the tick vector, Ixodes scapularis. Previous studies have characterized the tick transcriptome and proteome in response to A. phagocytophilum infection. However, in the post-genomic era, the integration of omics datasets through a systems biology approach allows network-based analyses to describe the complexity and functionality of biological systems such as host-pathogen interactions and the discovery of new targets for prevention and control of infectious diseases. This study reports for the first time a systems biology integration of metabolomics, transcriptomics and proteomics data to characterize essential metabolic pathways involved in the response of tick cells to A. phagocytophilum infection. The results showed that infection affected protein processing in endoplasmic reticulum and glucose metabolic pathways in tick cells. These results supported tick-Anaplasma co-evolution by providing new evidence of how tick cells limit pathogen infection, while the pathogen benefits from the tick cell response to establish infection. The results suggested that A. phagocytophilum induces protein misfolding to limit the tick cell response and facilitate infection, but requires protein degradation to prevent ER stress and cell apoptosis to survive in infected cells. Additionally, A. phagocytophilum may benefit from the tick cellâs ability to limit bacterial infection through PEPCK inhibition leading to decreased glucose metabolism, which also results in the inhibition of cell apoptosis that increases infection of tick cells. These results support the use of this experimental approach to systematically identify tick cell pathways and molecular mechanisms involved in tick-pathogen interactions Two samples with two replicates each were analyzed. Samples included Ixodes scapularis ISE6 cells uninfected (control) and infected with Anaplasma phagocytophilum human NY18 isolate.

Project description:Cellular senescence is a program of irreversible cell cycle arrest that normal cells undergo in response to progressive shortening of telomeres, changes in telomeric structure, oncogene activation or oxidative stress. The underlying signalling pathways, potentially of major clinicopathological relevance, are unknown. A major stumbling block to studying senescence has been the absence of suitable model systems because of the asynchrony of this process in heterogeneous cell populations. To simplify this process many investigators study oncogene-induced senescence due to expression of activated oncogenes where senescence occurs prematurely without telomere attrition and can be induced acutely in a variety of cell types. We have taken a different approach by making use of the finding that reconstitution of telomerase activity by introduction of the catalytic subunit of human telomerase alone is incapable of immortalising all human somatic cells, but inactivation of the p16-pRB and p53-p21 pathways are required in addition. The ability of SV40 large T antigen to inactivate the p16-pRB and p53-p21 pathways has enabled us to use a thermolabile mutant of LT antigen, in conjunction with hTERT, to develop conditionally immortalised human (HMF3A) fibroblasts that are immortal but undergo an irreversible growth arrest when the thermolabile LT antigen is inactivated leading to activation of pRB and p53. When these cells cease dividing, senescence-associated- b-galactosidase activity is induced and the growth-arrested cells have morphological features and express genes in common with senescent cells. Since these cells growth arrest in a synchronous manner they are an excellent starting point for dissecting the pathways that underlie cellular senescence and act downstream of p16-pRB and p53-p21 pathways. We have combined genome-wide expression profiling with genetic complementation to undertake identification of genes that are differentially expressed when these conditionally immortalised human fibroblasts undergo senescence upon activation of the p16-pRB and p53-p21 tumour suppressor pathways. Genes differentially expressed upon senescence will be identified by comparing arrays from growing versus senescent cells. Changes in gene expression due to the temperature shift will be eliminated by comparing with array data from the non-conditional HMF3S cells grown at 34°C ±0.5°C and 38°C ±0.5°C. To determine if the changes in gene expression upon senescence are specific and reversible, the set of differential genes will then be overlaid with array data from cells in which senescence has been bypassed by inactivation of the p16-pRB and p53-p21 tumour suppressor pathways

Project description:Cellular senescence is a program of irreversible cell cycle arrest that normal cells undergo in response to progressive shortening of telomeres, changes in telomeric structure, oncogene activation or oxidative stress. The underlying signalling pathways, potentially of major clinicopathological relevance, are unknown. A major stumbling block to studying senescence has been the absence of suitable model systems because of the asynchrony of this process in heterogeneous cell populations. To simplify this process many investigators study oncogene-induced senescence due to expression of activated oncogenes where senescence occurs prematurely without telomere attrition and can be induced acutely in a variety of cell types. We have taken a different approach by making use of the finding that reconstitution of telomerase activity by introduction of the catalytic subunit of human telomerase alone is incapable of immortalising all human somatic cells, but inactivation of the p16-pRB and p53-p21 pathways are required in addition. The ability of SV40 large T antigen to inactivate the p16-pRB and p53-p21 pathways has enabled us to use a thermolabile mutant of LT antigen, in conjunction with hTERT, to develop conditionally immortalised human (HMF3A) fibroblasts that are immortal but undergo an irreversible growth arrest when the thermolabile LT antigen is inactivated leading to activation of pRB and p53. When these cells cease dividing, senescence-associated- b-galactosidase activity is induced and the growth-arrested cells have morphological features and express genes in common with senescent cells. Since these cells growth arrest in a synchronous manner they are an excellent starting point for dissecting the pathways that underlie cellular senescence and act downstream of p16-pRB and p53-p21 pathways. We have combined genome-wide expression profiling with genetic complementation to undertake identification of genes that are differentially expressed when these conditionally immortalised human fibroblasts undergo senescence upon activation of the p16-pRB and p53-p21 tumour suppressor pathways.

			Action	DRS
		Other
	SRR17229500_1.fastq.gz	Fastqsanger.gz
	SRR17229500_2.fastq.gz	Fastqsanger.gz
	SRR17229501_1.fastq.gz	Fastqsanger.gz
	SRR17229501_2.fastq.gz	Fastqsanger.gz

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Dissecting molecular pathways of neuropathology of PHTS by transcriptomics and proteomics

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