Autophagy Activation Clears ELAVL1/HuR-Mediated Accumulation of SQSTM1/p62 during Proteasomal Inhibition in Human Retinal Pigment Epithelial Cells

Abstract

Age-related macular degeneration (AMD) is the most common reason of visual impairment in the elderly in the Western countries. The degeneration of retinal pigment epithelial cells (RPE) causes secondarily adverse effects on neural retina leading to visual loss. The aging characteristics of the RPE involve lysosomal accumulation of lipofuscin and extracellular protein aggregates called “drusen”. Molecular mechanisms behind protein aggregations are weakly understood. There is intriguing evidence suggesting that protein SQSTM1/p62, together with autophagy, has a role in the pathology of different degenerative diseases. It appears that SQSTM1/p62 is a connecting link between autophagy and proteasome mediated proteolysis, and expressed strongly under the exposure to various oxidative stimuli and proteasomal inhibition. ELAVL1/HuR protein is a post-transcriptional factor, which acts mainly as a positive regulator of gene expression by binding to specific mRNAs whose corresponding proteins are fundamental for key cellular functions. We here show that, under proteasomal inhibitor MG-132, ELAVL1/HuR is up-regulated at both mRNA and protein levels, and that this protein binds and post-transcriptionally regulates SQSTM1/p62 mRNA in ARPE-19 cell line. Furthermore, we observed that proteasomal inhibition caused accumulation of SQSTM1/p62 bound irreversibly to perinuclear protein aggregates. The addition of the AMPK activator AICAR was pro-survival and promoted cleansing by autophagy of the former complex, but not of the ELAVL1/HuR accumulation, indeed suggesting that SQSTM1/p62 is decreased through autophagy-mediated degradation, while ELAVL1/HuR through the proteasomal pathway. Interestingly, when compared to human controls, AMD donor samples show strong SQSTM1/p62 rather than ELAVL1/HuR accumulation in the drusen rich macular area suggesting impaired autophagy in the pathology of AMD.

Crosstalk between autophagy and proteasome protein degradation systems: possible implications for cancer therapy

“…Aggresomes are predominantly present in perinuclear compartments, and they are considered to be a place of selective autophagy of damaged proteins that are isolated from other cytoplasmic components [129]. The clearance of polyubiquitinated proteins present in aggresome is mediated by ubiquitin-binding proteins like p62/SQSTM1 (Figure 2 to be a connecting link between autophagy and proteasome-mediated proteolysis, and its level strongly increases during exposure to various oxidative agents and proteasomal inhibitors, such as MG-132, lactacystin, epoxomicin and PSI [66,[130][131][132]. Furthermore, proteasomal inhibition caused by MG-132 leads to accumulation of p62/SQSTM1 bound irreversibly to perinuclear protein aggregates [132].…”

“…The clearance of polyubiquitinated proteins present in aggresome is mediated by ubiquitin-binding proteins like p62/SQSTM1 (Figure 2 to be a connecting link between autophagy and proteasome-mediated proteolysis, and its level strongly increases during exposure to various oxidative agents and proteasomal inhibitors, such as MG-132, lactacystin, epoxomicin and PSI [66,[130][131][132]. Furthermore, proteasomal inhibition caused by MG-132 leads to accumulation of p62/SQSTM1 bound irreversibly to perinuclear protein aggregates [132]. This phenomenon may be explained, at least partially, on the basis of experiments performed with MG-132 in the ARPE-19 cell line [132].…”

“…Furthermore, proteasomal inhibition caused by MG-132 leads to accumulation of p62/SQSTM1 bound irreversibly to perinuclear protein aggregates [132]. This phenomenon may be explained, at least partially, on the basis of experiments performed with MG-132 in the ARPE-19 cell line [132]. Viiri et al studied the involvement of ELAVL1/HuR protein in the regulation of p62/ /SQSTM1 synthesis.…”

“…ELAVL1/HuR is a post-transcriptional factor, which acts mainly as a positive regulator of gene expression by binding to specific mRNAs, which corresponding proteins play a key role in cell functions. During proteasomal inhibition ELAVL1/HuR was up-regulated at both mRNA and protein levels, and was shown to bind with and post-transcriptionally regulate p62/SQSTM1 mRNA [132]. On the other hand, substantial decreases in p62/SQSTM1 levels after proteasomal inhibition in human osteosarcoma (HOS) cells have been reported [77].…”

Induction of VEGFA mRNA translation by CoCl2 mediated by HuR

“…15 Indeed, HuR promotes the expression of general stress-response proteins (HSP70, HO-1, SOD1, p62) and hypoxia-response proteins, including HIF-1a and VEGFA. [16][17][18][19][20] Through its influence on subsets of cellular proteins, HuR has been linked to various pathologies, including inflammatory diseases and cancer. 12,13 With respect to diabetic retinopathy, the levels of both HuR and VEGFA have been shown to be altered in both in vitro and in vivo models.…”