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A comparative kidney transcriptome analysis of bicarbonate-loaded insrr-null mice

ABSTRACT: The maintenance of physiological pH level is critical for the life of all organisms. Well known, that kidney is key organ for acid-base regulation in vertebrates. Metabolic alkalosis is the most common acid-base disorder in hospitalized patients, particularly in the surgical critical care unit. The receptor tyrosine kinase IRR (insulin receptor-related receptor), a receptor like the insulin receptor, is involved in acid-base regulation by kidney due to its ability to autophosphorylate under alkalization of extracellular medium (pH > 7.9). IRR expressed in only β-intercalated cells. In mice with a knockout of the insrr gene, which encoding IRR, bicarbonate secretion to the urine is impaired. Here for the first time, we present analysis of kidney transcriptomes from wild type and insrr gene knockout mice under basal or bicarbonate-loaded conditions by 250 mM NaHCO3 in drinking water. Analysis of NGS sequencing data revealed changes in the expression of a number of genes in the kidney. Using real-time PCR we confirmed change of expression of the slc51b, slc26a4, rps7, slc5a2, aqp6, plcd1, gapdh, rny3, lypd2, kcnk5, slc6a6 , atp6v1g3 genes in IRR knockout mice. Also, we found that expression of kcnk5 gene is increased in wild type mice after bicarbonate loading but didn’t change in knockout mice. Gene set enrichment between the IRR knockout and wildtype samples identified that knockout causes alterations in expression of genes related mostly to the ATP metabolic and electron transport chain processes. Thus, for the first time, we described some details of the functioning of the IRR in the kidneys.

ORGANISM(S): Mus musculus

PROVIDER: GSE200638 | GEO | 2022/04/13

REPOSITORIES: GEO

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Project description:Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown with aeration to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Analysis of variance with a significance level of 0.001 resulted in 98% power to detect genes showing a twofold difference in expression. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High (highest expression at pH 5.0), Acid Low (lowest expression at pH 5.0), Base High (highest at pH 8.7), Base Low (lowest at pH 8.7), Neutral High (highest at pH 7.0, lower in acid or base), and Neutral Low (lowest at pH 7.0, higher at both pH extremes). Flagellar and chemotaxis genes were repressed at pH 8.7 (Base Low cluster), where the cell's transmembrane proton potential is diminished by the maintenance of an inverted pH gradient. High pH also repressed the proton pumps cytochrome o (cyo) and NADH dehydrogenases I and II. By contrast, the proton-importing ATP synthase F1Fo and the microaerophilic cytochrome d (cyd), which minimizes proton export, were induced at pH 8.7. These observations are consistent with a model in which high pH represses synthesis of flagella, which expend proton motive force, while stepping up electron transport and ATPase components that keep protons inside the cell. Acid-induced genes, on the other hand, were coinduced by conditions associated with increased metabolic rate, such as oxidative stress. All six pH-dependent clusters included envelope and periplasmic proteins, which directly experience external pH. Overall, this study showed that (i) low pH accelerates acid consumption and proton export, while coinducing oxidative stress and heat shock regulons; (ii) high pH accelerates proton import, while repressing the energy-expensive flagellar and chemotaxis regulons; and (iii) pH differentially regulates a large number of periplasmic and envelope proteins. Keywords: Steady State

Project description:Kidneys are essential for acid-base homeostasis, especially when organisms cope with changes in acid or base dietary intake. Because collecting ducts constitute the final site for regulating urine acid-base balance, we undertook to identify the gene network involved in acid-base transport and regulation in the mouse outer medullary collecting duct (OMCD). For this purpose, we combined kidney functional studies and quantitative analysis of gene expression in OMCDs, by transcriptome and candidate gene approaches, during metabolic acidosis. Furthermore, to better delineate the set of genes concerned with acid-base disturbance, the OMCD transcriptome of acidotic mice was both compared to that of normal mice and mice undergoing an adaptative response through potassium depletion. Metabolic acidosis, achieved through a NH4Cl-supplemented diet for 3 days, not only induced acid secretion, but also stimulated the aldosterone and vasopressin systems and triggered cell proliferation. Accordingly, metabolic acidosis increased the expression of genes involved in acid-base transport, sodium transport, water transport and cell proliferation. In particular, more than 25 transcripts encoding proteins involved in urine acidification (subunits of H-ATPase, kidney anion exchanger, chloride channel Clcka, carbonic anhydrase 2, aldolase) were co-regulated during acidosis. We propose that these trancripts cooperating to a same function and co-regulated during acidosis constitute a functional regulon. Keywords: SAGE; Kidney collecting duct; Mouse; Metabolic acidosis; V-ATPase Approximately 650 OMCDs were microdissected, as previously described from collagenase-treated kidneys of 6 mice fed the NH4Cl-supplemented diet for 3 days. 20960 tags were sequenced in the acidosis SAGE library. After withdrawing duplicate ditags and linker sequences, the 19146 tags remaining corresponded to 8847 different tags.This acidosis library is similar in size and diversity to the normal mouse OMCD libraries previously generated (GEO accession number : GSM23478). The tag abundance in the two library was compared and discussed.

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A comparative kidney transcriptome analysis of bicarbonate-loaded insrr-null mice

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