Inactivation of Tristetraprolin in Chronic Hypoxia Provokes the Expression of Cathepsin B

Abstract

bMacrophages play important roles in many diseases and are frequently found in hypoxic areas. A chronic hypoxic microenvironment alters global cellular protein expression, but molecular details remain poorly understood. Although hypoxiainducible factor (HIF) is an established transcription factor allowing adaption to acute hypoxia, responses to chronic hypoxia are more complex. Based on a two-dimensional differential gel electrophoresis (2D-DIGE) approach, we aimed to identify proteins that are exclusively expressed under chronic but not acute hypoxia (1% O 2 ). One of the identified proteins was cathepsin B (CTSB), and a knockdown of either HIF-1␣ or -2␣ in primary human macrophages pointed to an HIF-2␣ dependency. Although chromatin immunoprecipitation (ChIP) experiments confirmed HIF-2 binding to a CTSB enhancer in acute hypoxia, an increase of CTSB mRNA was evident only under chronic hypoxia. Along those lines, CTSB mRNA stability increased at 48 h but not at 8 h of hypoxia. However, RNA stability at 8 h of hypoxia was enhanced by a knockdown of tristetraprolin (TTP). Inactivation of TTP under prolonged hypoxia was facilitated by c-Jun N-terminal kinase (JNK), and inhibition of this kinase lowered CTSB mRNA levels and stability. We postulate a TTP-dependent mechanism to explain delayed expression of CTSB under chronic hypoxia.C hronic diseases such as diabetes, atherosclerosis, and cancer are characterized by hypoxic areas resulting, for example, from compromised perfusion of narrowed or leaky vessels. Cells of the immune system are involved in the outcome of these diseases. As part of the innate immune system, macrophages actively regulate inflammation but also the resolution of inflammation as well as tissue regeneration and remodeling. Macrophages invade hypoxic areas attracted by a number of cytokines produced by hypoxic cells. To survive and operate in a hypoxic environment, cells need a variety of adaptive mechanisms (1, 2).Hypoxia-inducible factors (HIFs) are important to coordinate hypoxic responses and consist of a constitutively expressed ␤-subunit and an oxygen-regulated ␣-subunit. Both are members of the helix-loop-helix/Per, ARNT, and SIM (PAS) transcription factor family (1, 3). Among the ␣-subunits, HIF-1␣ and HIF-2␣ are best characterized. Both contain an oxygen-dependent degradation domain (ODD) with two conserved prolyl residues that are hydroxylated by prolyl hydroxylases (PHDs) 1 to 3 when sufficient oxygen is available, allowing their proteasomal degradation (4, 5). PHDs are impaired under hypoxia, which in turn causes accumulation and translocation of HIF-␣ into the nucleus. The ␣-subunit forms a heterodimer with the ␤-subunit and binds to hypoxiaresponsive elements (HRE) in regulatory regions of target genes (6). By recruiting cofactors like p300 or CBP, the HIF proteins enhance transcription of about 400 target genes (7,8). Although HIF abundance is mostly regulated by protein stability, regulation of HIF-1␣ mRNA via binding of tristetraprolin (TTP) to AU-rich elements (AREs) in the 3...

Chronic Hypoxia Enhances β-Oxidation-Dependent Electron Transport via Electron Transferring Flavoproteins

“…During the progression from acute to chronic hypoxia, we noticed a decline of HIF-1α, HIF-2α, as well as HIF-target gene expression [5,22]. Reaching this new ‘set point’, which is apparently distinct from acute hypoxic responses, cells may reset their metabolism accordingly.…”

Identification of the Cysteine Protease Legumain as a Potential Chronic Hypoxia-Specific Multiple Myeloma Target Gene

“…Moreover, unlike solid tumor cells, liquid tumors, including MM, are able to translocate when confronted with unfavorable oxygen conditions, e.g., too low for physiological function [ 43 ]. Therefore, to analyze how chronic hypoxia affect MM cells in vitro, four MM cell lines (RPMI-8226, LP-1, OPM-2 and KMS-12-BM) were adapted to physiological oxygen concentration in vitro (1% O 2 and 5% CO 2 ) over a period of 7 days and subsequently tested for the mRNA expression of predefined marker genes for acute hypoxia (Egl-9 family hypoxia inducible factor 1, EGLN1 ( PHD2 ); adrenomedullin, ADM ) and chronic hypoxia (H4 clustered histone 1, H4C1 ; osteoclast stimulating factor 1, OSTF1 ) [ 44 , 45 ] ( Figure 1 ). For the expression of the acute hypoxia marker EGLN1 , a significant increase was detected up to day 3, which regressed during the course of the experiment and reached the expression level of day 0 after 7 days of hypoxia ( Figure 1 A).…”

Strap associates with Csde1 and affects expression of select Csde1-bound transcripts

“…Thoc5 is a nuclear exporter of mRNAs essential for the maintenance of hematopoiesis [ 44 ]. Hmbs catalyzes a crucial step in heme biosynthesis and is oppositely regulated from Hif-1 under hypoxic conditions [ 36 ], and Ostf-1 is induced by hypoxia [ 37 , 45 ]. Regulation of these transcripts suggest a possible role for Strap in the regulation of terminal erythroid differentiation and the hypoxic response.…”