GATA6-AS1 Regulates Intestinal Epithelial Mitochondrial Functions, and its Reduced Expression is Linked to Intestinal Inflammation and Less Favourable Disease Course in Ulcerative Colitis.

"rna sequencing [rnaseq] data from protect [gse109142] seem [gse159495] risk [rectal gse117993 ileal gse101794] and coeliac cohort [prjna528755[21]] were previously published and are available with the indicated accessions and source [gse199906] data were deposited in geo.; data availability rna sequencing [rnaseq] data from protect [gse109142] seem [gse159495] risk [rectal gse117993 ileal gse101794] and coeliac cohort [prjna528755[21]] were previously published and are available with the indicated accessions and source [gse199906] data were deposited in geo. we performed co-expression analyses within protect followed by functional annotation enrichment of the 684 co-expressed and better annotated protein coding genes [ figure 1e dataset s1].; lines indicate median and upper and lower quartile [c] gata6-as1 is further reduced in more severe uc cases based on clinical [pucai] combined clinical-endoscopic [total mayo score] and endoscopic disease severity [endoscopic mayo score] in the protect cohort [d] gata6-as1 mrna reduction is linked to less favourable disease outcome [w4r or w52sfr] in the protect cohort gata6-as1 levels in controls are given as a reference [e] functional annotation enrichments using toppgene/toppcluster and cytoscape of the gata6-as1 co-expression network in protect using euclidean distance [full list and fdr values are given in supplementary dataset s1 ].; functional annotation of these proteins indicated strong and significant enrichments for metabolic processes mitochondria and tca cycle [ figure 3a-c and supplementary dataset s2 ] which was consistent with the gata6-as1 co-expression in patients which was also significantly enriched for metabolic and mitochondrial functions [ figure 1e ; supplementary dataset s1 ].; binding of gata6-as1 rna was confirmed by pcr [b c] the gata6-as1 interactome in differentiated caco-2 cells identified 285 proteins using mass spectrometry [ t -test: delta lfq >= 0 1 and fdr <= 0 05; supplementary dataset s2 ] [b] heatmap of gata6-as1 top curated interacting targets identified by mass spectrometry; green and red colours indicate low and high binding respectively; ctl columns are binding to non-specific control oligos [c] functional enrichments of the 285 protein targets indicated enrichment of mitochondrial functions [toppgene/toppcluster ] [d] gata6-as1 silencing [sh1 sh2] was compared to scrambled non-specific shrna [ctl] using qrt-pcr [normalized to gapdh and to controls] [e] jc1 staining and facs analysis were used to define the mitochondrial membrane potential [mmp]; schematic representation of mmp calculated as the ratio of red/green fluorescence.; seventy-four differentially expressed metabolites were identified in cells and 49 in their media [paired rank mean test and fdr < 0 1 supplementary dataset s3 ] with a prominent reduction in the tca cycle metabolites alpha-ketoglutarate [akg] and malate and in associated metabolites including glutamate aspartate and n -acetyl-aspartate [naa figure 4b ] upon gata6-as1 knockdown.; [a] schematic representation of the experiment and pcoa plot of the extracted metabolites based on canberra distance indicating differences by both gata6-as1 expression and by treatment [b] heatmap representing significantly different metabolites in cells [left heatmap] and their media [right heatmap rank-mean test and fdr < 0 1] between gata6-as1 knockdown [sh1 and sh2] and control cells with visualization of the metabolites in both treated and untreated cells [full comparisons in supplementary dataset s3 ]. data availability rna sequencing [rnaseq] data from protect [gse109142"

"Conflict of Interest The following authors have no conflicts of interest: KES, TB, AA, MB, KLV, NL, HAE, KB, RBY, SAA, SRM, SK, IA, EGS, BW, IB, EG, TG, SBH, IU, LAD, YH. JSH: Advisory Board Janssen, consultant: Takeda, Pfizer, Bristol Myers Squibb, Boehringer Ingleheim, and Eli Lilly."

"Funding This work was supported by the ERC starting grant [YH, grant No. 758313], the Israel Science Foundation [YH, grant No. 908/15], the I-CORE programme [YH, grant No. 41/11], the Helmsley Charitable Trust, and NIDDK P30 DK078392 [Integrative Morphology and Gene Expression Cores]. PROTECT was supported by the NIDDK 5U01DK095745, RISK was supported by Crohn’s & Colitis Foundation, SEEM by the Bill and Melinda Gates Foundation [OPP1144149 and OPP1138727], and SOURCE is supported by the Helmsley Charitable Trust. The funding sources did not play a role in writing of the manuscript or the decision to submit it for publication, did not play a role in data collection, analysis or interpretation; trial design; patient recruitment; or any aspect pertinent to the study."