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Transcriptomic changes triggered by hypoxia: evidence for HIF-1α-independent, [Na+]i/[K+]i-mediated, excitation-transcription coupling.

Koltsova SV, Shilov B, Birulina JG, Akimova OA, Haloui M, Kapilevich LV, Gusakova SV, Tremblay J, Hamet P, Orlov SN - PLoS ONE (2014)

Bottom Line: Using global gene expression profiling we found that Na+,K+-ATPase inhibition by ouabain or K+-free medium in rat aortic vascular smooth muscle cells (RASMC) led to the differential expression of dozens of genes whose altered expression was previously detected in cells subjected to hypoxia and ischemia/reperfusion.In ouabain-treated RASMC, low-Na+, high-K+ medium abolished amplification of the [Na+]i/[K+]i ratio as well as the increased expression of all tested genes.In cells subjected to hypoxia and glucose deprivation, dissipation of the transmembrane gradient of Na+ and K+ completely eliminated increment of Fos, Atf3, Ptgs2 and Per2 mRNAs and sharply diminished augmentation expression of Klf10, Edn1, Nr4a1 and Hes1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Moscow State University, Moscow, Russia; Department of Medicine, Centre de recherche, Centre hospitalier de l'Université de Montréal, Montreal, Quebec, Canada.

ABSTRACT
This study examines the relative impact of canonical hypoxia-inducible factor-1alpha- (HIF-1α and Na+i/K+i-mediated signaling on transcriptomic changes evoked by hypoxia and glucose deprivation. Incubation of RASMC in ischemic conditions resulted in ∼3-fold elevation of [Na+]i and 2-fold reduction of [K+]i. Using global gene expression profiling we found that Na+,K+-ATPase inhibition by ouabain or K+-free medium in rat aortic vascular smooth muscle cells (RASMC) led to the differential expression of dozens of genes whose altered expression was previously detected in cells subjected to hypoxia and ischemia/reperfusion. For further investigations, we selected Cyp1a1, Fos, Atf3, Klf10, Ptgs2, Nr4a1, Per2 and Hes1, i.e. genes possessing the highest increments of expression under sustained Na+,K+-ATPase inhibition and whose implication in the pathogenesis of hypoxia was proved in previous studies. In ouabain-treated RASMC, low-Na+, high-K+ medium abolished amplification of the [Na+]i/[K+]i ratio as well as the increased expression of all tested genes. In cells subjected to hypoxia and glucose deprivation, dissipation of the transmembrane gradient of Na+ and K+ completely eliminated increment of Fos, Atf3, Ptgs2 and Per2 mRNAs and sharply diminished augmentation expression of Klf10, Edn1, Nr4a1 and Hes1. In contrast to low-Na+, high-K+ medium, RASMC transfection with Hif-1a siRNA attenuated increments of Vegfa, Edn1, Klf10 and Nr4a1 mRNAs triggered by hypoxia but did not impact Fos, Atf3, Ptgs2 and Per2 expression. Thus, our investigation demonstrates, for the first time, that Na+i/K+i-mediated, Hif-1α- -independent excitation-transcription coupling contributes to transcriptomic changes evoked in RASMC by hypoxia and glucose deprivation.

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Position of (A/G)CGTG consensus within 10,000 bp 5′-UTR of genes listed in Table 5.
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pone-0110597-g008: Position of (A/G)CGTG consensus within 10,000 bp 5′-UTR of genes listed in Table 5.

Mentions: Several research teams reported that HIF-1α regulates gene expression in ischemic tissues via interaction of HIF-1α/HIF-1β heterodimer with HREs containing (A/G)CGTG consensus in promoter/enhancer regions of the target gene's DNA such as VEGFA [45] and EDN1 [46]. Considering this, we employed SCOPE service (Suite for Computational Identification of Promoter Elements): http://genie.dartmouth.edu/scope/[47] for the search of (A/G)CGTG consensus within 5′-untranslated regions (5′-UTR) of [Na+]i/[K+]i-sensitive genes listed in Table 4. Using this approach we found numerous (A/G)CGTG sequences within 5′-UTR encoding canonical HIF-1-sensitve genes (Edn1 and Vegfa) as well as all [Na+]i/[K+]i-sensitive genes listed in Table 4. Importantly, we failed to find any fixed position for this consensus within 10,000 bp 5′-UTRs of HIF1α-sensitive vs HIF1α-resistant genes (Fig. 8). Moreover, we observed that in several [Na+]i/[K+]i-sensitive genes proximal 1,500 bp segments of 5′-UTRs are more abundant with (A/G)CGTG sequence as compared to canonical HIF-sensitive transcripts (Fig. 9). Thus, 1,500 bp 5′-UTRs of Atf3 and Edn1 contains 8 and 3 (A/G)CGTG sequences. This observation is also confirmed by Sig Value parameter having a value of 28.4 for 1500 bp 5′-UTRs of genes listed in Table 5. If the search is not restricted to positions of 1500 bp, Sig Value is negative indicating the absence of predictive capabilities for the consensus sequence.


Transcriptomic changes triggered by hypoxia: evidence for HIF-1α-independent, [Na+]i/[K+]i-mediated, excitation-transcription coupling.

Koltsova SV, Shilov B, Birulina JG, Akimova OA, Haloui M, Kapilevich LV, Gusakova SV, Tremblay J, Hamet P, Orlov SN - PLoS ONE (2014)

Position of (A/G)CGTG consensus within 10,000 bp 5′-UTR of genes listed in Table 5.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4222758&req=5

pone-0110597-g008: Position of (A/G)CGTG consensus within 10,000 bp 5′-UTR of genes listed in Table 5.
Mentions: Several research teams reported that HIF-1α regulates gene expression in ischemic tissues via interaction of HIF-1α/HIF-1β heterodimer with HREs containing (A/G)CGTG consensus in promoter/enhancer regions of the target gene's DNA such as VEGFA [45] and EDN1 [46]. Considering this, we employed SCOPE service (Suite for Computational Identification of Promoter Elements): http://genie.dartmouth.edu/scope/[47] for the search of (A/G)CGTG consensus within 5′-untranslated regions (5′-UTR) of [Na+]i/[K+]i-sensitive genes listed in Table 4. Using this approach we found numerous (A/G)CGTG sequences within 5′-UTR encoding canonical HIF-1-sensitve genes (Edn1 and Vegfa) as well as all [Na+]i/[K+]i-sensitive genes listed in Table 4. Importantly, we failed to find any fixed position for this consensus within 10,000 bp 5′-UTRs of HIF1α-sensitive vs HIF1α-resistant genes (Fig. 8). Moreover, we observed that in several [Na+]i/[K+]i-sensitive genes proximal 1,500 bp segments of 5′-UTRs are more abundant with (A/G)CGTG sequence as compared to canonical HIF-sensitive transcripts (Fig. 9). Thus, 1,500 bp 5′-UTRs of Atf3 and Edn1 contains 8 and 3 (A/G)CGTG sequences. This observation is also confirmed by Sig Value parameter having a value of 28.4 for 1500 bp 5′-UTRs of genes listed in Table 5. If the search is not restricted to positions of 1500 bp, Sig Value is negative indicating the absence of predictive capabilities for the consensus sequence.

Bottom Line: Using global gene expression profiling we found that Na+,K+-ATPase inhibition by ouabain or K+-free medium in rat aortic vascular smooth muscle cells (RASMC) led to the differential expression of dozens of genes whose altered expression was previously detected in cells subjected to hypoxia and ischemia/reperfusion.In ouabain-treated RASMC, low-Na+, high-K+ medium abolished amplification of the [Na+]i/[K+]i ratio as well as the increased expression of all tested genes.In cells subjected to hypoxia and glucose deprivation, dissipation of the transmembrane gradient of Na+ and K+ completely eliminated increment of Fos, Atf3, Ptgs2 and Per2 mRNAs and sharply diminished augmentation expression of Klf10, Edn1, Nr4a1 and Hes1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Moscow State University, Moscow, Russia; Department of Medicine, Centre de recherche, Centre hospitalier de l'Université de Montréal, Montreal, Quebec, Canada.

ABSTRACT
This study examines the relative impact of canonical hypoxia-inducible factor-1alpha- (HIF-1α and Na+i/K+i-mediated signaling on transcriptomic changes evoked by hypoxia and glucose deprivation. Incubation of RASMC in ischemic conditions resulted in ∼3-fold elevation of [Na+]i and 2-fold reduction of [K+]i. Using global gene expression profiling we found that Na+,K+-ATPase inhibition by ouabain or K+-free medium in rat aortic vascular smooth muscle cells (RASMC) led to the differential expression of dozens of genes whose altered expression was previously detected in cells subjected to hypoxia and ischemia/reperfusion. For further investigations, we selected Cyp1a1, Fos, Atf3, Klf10, Ptgs2, Nr4a1, Per2 and Hes1, i.e. genes possessing the highest increments of expression under sustained Na+,K+-ATPase inhibition and whose implication in the pathogenesis of hypoxia was proved in previous studies. In ouabain-treated RASMC, low-Na+, high-K+ medium abolished amplification of the [Na+]i/[K+]i ratio as well as the increased expression of all tested genes. In cells subjected to hypoxia and glucose deprivation, dissipation of the transmembrane gradient of Na+ and K+ completely eliminated increment of Fos, Atf3, Ptgs2 and Per2 mRNAs and sharply diminished augmentation expression of Klf10, Edn1, Nr4a1 and Hes1. In contrast to low-Na+, high-K+ medium, RASMC transfection with Hif-1a siRNA attenuated increments of Vegfa, Edn1, Klf10 and Nr4a1 mRNAs triggered by hypoxia but did not impact Fos, Atf3, Ptgs2 and Per2 expression. Thus, our investigation demonstrates, for the first time, that Na+i/K+i-mediated, Hif-1α- -independent excitation-transcription coupling contributes to transcriptomic changes evoked in RASMC by hypoxia and glucose deprivation.

Show MeSH
Related in: MedlinePlus