HSP47: Regulation

ICC of Heat Shocked HeLa cells
ICC staining of heat shocked HeLa cells using Anti-Hsp47 (clone: 1C4-1A6)

When cells are subjected to environmental stress, they respond by enhancing expression of HSPs. The rapid induction of HSP in response to environmental stress is based on a variety of genetic and biochemical processes referred to as the heat shock response (HSR) 169. HSR is preferentially regulated at the transcription level by heat shock factors (HSF). Among them, HSF-1 is considered as being the key transcription factor of stress-inducible HSPs 170. Under normal conditions, HSF-1 exists as an inactive monomer in the cytosol in conjunction with further chaperones such as Hsp70 and Hsp90 171. Exposure to stress induces the release of Hsp70 and Hsp90 proteins fom the complex with subsequent formation of HSF-1 homotrimers capable of binding to heat shock elements (HSEs) up-stream of HSP promoters, thereby triggering HSP gene transcription 172, 173. Hsp47 is encoded by a single copy gene in the human genome (SERPINH1) 37 as well as in the mouse genome (Serpinh1) 38. The mouse Serpinh1 gene harbors six exons and five introns, a promoter region bearing a TATA box, binding sites for specificity protein 1 (Sp-1), transcription factor activator protein 1 (AP-1), retinoic acid-responsive elements (RAREs) and one heat shock element (HSE) 38, 174. In contrast, the human SERPINH1 gene contains five exons and lacks binding sites for the transcription factors Sp-1 and AP-1, respectively 38, 175. Hsp47 was originally characterized as a 47 kDa procollagen-binding protein whose expression is up-regulated upon heat shock 15. SERPINH1 is transcriptionally regulated by HSF-1 and has been identified to be induced by both, TGF-β and interleukins 176. The heat inducibility of Hsp47 is mediated by one HSE located 180 bp up-stream of the transcription start 28, 38, 177. Upon cellular stress, trimeric HSF-1 binds to HSE and promotes the transcription of SERPINH1. As outlined before, the expression of Hsp47 varies with tissue type, developmental stage and stress conditions. Notwithstanding its heat inducibility, Hsp47 is constitutively expressed under normal conditions and Hsp47 expression levels have been shown to correlate with those of collagens in numerous tissues and cell types. Two regions in the promoter region of SERPINH1 seem to play a pivotal role in regulating the cell-type and tissue-specific expression of Hsp47. The most distal GC box in the promoter region and two additional elements (BS5-B in the first intron and EP7-D in the second intron) have been reported as being responsible for basic expression and tissue-specific expression of Hsp47 (Serpin H1), respectively. While constitutive Hsp47 expression demands an Sp-1 binding site 280 bp up-stream of the transcription start, tissue-specific expression relies on the binding of Sp-1/Sp-2 and KLF-6/Zf9 to BS5-B in the first intron and most probably to EP7-D in the second intron 2. It is interesting to note that the sequence element BS5-B in the first intron acts as an enhancer element contributing to tissue-specific Hsp47 expression, since both transcription factors, Sp-1 and Zf9 have been noted to enhance the synthesis of collagen type I 2, 178.

Data raised by the group of Yoriko Atomi provided evidence for the relation of Hsp47 expression to gravity changes. The results clearly demonstrated that low or zero gravity causes a marked down-regulation of Hsp47 whereas hypergravity induces an up-regulation of Hsp47 179. These findings should be viewed in the context of skeletal muscle unloading induced by spaceflight or bed rest that both can lead to muscle atrophy. The clear Hsp47 down-regulation under lower gravity might be attributed to osteoporosis emerging from microgravity conditions as occurring during longterm space missions.

Apart from its transcriptional regulation, Hsp47 protein levels have also been found as being regulated at the post-transcriptional level. In this context, the discovery of micro-RNA (miRNA) identified this RNA subtype as a crucial player in regulating translation of many genes. miRNAs are a class of small non-coding RNAs that negatively regulate gene expression by binding to target mRNAs. Global alterations in miRNAs can be observed in a number of disease states including cancer 180-182. Several investigations identified miR-29 which binds to the 3’-untranslated region in the SERPINH1 mRNA 167. In human tumor tissues, the expression of Hsp47 was found as being inversely associated with the expression of miR-29 167, 183-187. The miR-29 family has been implicated in the regulation of ECM genes, suggesting miR-29 markedly suppresses cancer cell migration and invasiveness. Ectopic expression of miR-29b or silencing of Hsp47 was noted to suppress malignant phenotypes of breast cancer cells by reducing deposition of collagen and fibronectin 167. In addition, Hsp47 up-regulation and miR-29 down-regulation were associated with poor survival outcomes in breast cancer patients revealing that up-regulated Hsp47 expression promotes cancer progression at least in part by boosting deposition of ECM proteins 167.

Similar to many other chaperones, Hsp47 is a phosphoprotein 30 whose expression and function can be further modulated by co- and post-translational modifications such as N-glycosylation, succinylation, and acetylation 31-33. GlcNAc oligosaccharide sidechains in Hsp47 have been proposed to represent the structural requirements for the intracellular interaction of Hsp47 with the calcium/phospholipid-binding protein annexin V 31-33. To what extend phosphorylation especially affects the molecular or biological functions of the chaperone is still unclear. A previous study revealed that SERPINH1 is silenced by aberrant methylation of promoter CpG islands in a number of different tumor types, and that Hsp47 deficiency results in enhanced levels of collagen I and IV 188. In endothelial cells, K165 could be identified as the primary acetylation site in Hsp47 32. Since HSPs have been reported to be crucially involved in modulating statin-derived endothelial homeostasis 189, post-translational acetylation might be implicated in the early regulatory functions of these HSPs including Hsp47. However, further investigations are warranted in order to support this hypothesis.