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PRKACA_DnaJ overexpression in HEK293 cells Phosphoproteomics

ABSTRACT: Phosphoproteomics studies of PKA catalytic subunit or its DNAJB1-fused chimeric form overexpressed in HEK293 cells, under various conditions including inhibitors. Also includes in vitro rephosphorylation of cell lysates by each construct.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Laurie Parker

PROVIDER: MSV000086707 | MassIVE | Fri Jan 15 13:22:00 GMT 2021

SECONDARY ACCESSION(S): PXD023638

REPOSITORIES: MassIVE

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Project description:GSM1056261-GSM1056272: NFX1-123 has been shown to associate with a number of RNA processing proteins, such as cytoplasmic poly(A) binding proteins (PABPC), to affect mRNA stability and translational efficiency of target genes. The high-risk human papillomavirus type 16E6 (HPV 16E6) induces telomerase activity by activation of hTERT, the catalytic subunit of telomerase. NFX1-123 can bind to hTERT mRNA and increase its stability in HPV 16E6 expressing keratinocytes (16E6 NHKs). Little is known regarding what other transcripts, and downstream signaling pathways, may be dependent on NFX1-123. In order to determine additional cellular transcripts affected by HPV 16E6 and NFX1-123, we assessed global gene expression changes in cells in which NFX1-123 was overexpressed or knocked down by short hairpin RNAs (shRNA) when compared to an isogenic scramble control, in three independently derived 16E6 NHKs GSM1212499-GSM1212510: NFX1-123 has been shown to associate with a number of RNA processing proteins, such as cytoplasmic poly(A) binding proteins (PABPC), to affect mRNA stability and translational efficiency of target genes. The high-risk human papillomavirus type 16E6 (HPV 16E6) induces telomerase activity by activation of hTERT, the catalytic subunit of telomerase. NFX1-123 can bind to hTERT mRNA and increase its stability in HPV 16E6 expressing keratinocytes (16E6 NHKs). Little is known regarding what other transcripts, and downstream signaling pathways, may be dependent on NFX1-123. In order to determine additional cellular transcripts affected solely by NFX1-123, we assessed global gene expression changes in cells in which NFX1-123 was overexpressed or knocked down by short hairpin RNAs (shRNA) when compared to an isogenic scramble control, in three independently derived NHKs. GSM1056261-GSM1056272: Three independent 16E6 expressing NHK cell lines were expanded and transduced with: short hairpin RNA (sh1) that knocked down NFX1-123 by 20%, short hairpin RNA (sh3) that knocked down NFX1-123 by 80%; a non-targeting isogenic shRNA scramble control; or a NFX1-123 overexpression construct with a FLAG-tag (FNFX1-123WT) that increased its RNA expression on average 3.5-fold. In total, twelve samples were used for the microarray, derived from the three initial NHK cell lines. GSM1212499-GSM1212510: Three independent NHK cell lines were expanded and transduced with: short hairpin RNA (sh1) that knocked down NFX1-123 by 40%, short hairpin RNA (sh3) that knocked down NFX1-123 by 83%; a non-targeting isogenic shRNA scramble control; or a NFX1-123 overexpression construct with a FLAG-tag (FNFX1-123WT) that increased its RNA expression on average 14.0-fold. In total, twelve samples were used for the microarray, derived from the three initial NHK cell lines.

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PRKACA_DnaJ overexpression in HEK293 cells Phosphoproteomics

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