4-PBA ameliorates cellular homeostasis in fibroblasts from osteogenesis imperfecta patients by enhancing autophagy and stimulating protein secretion

Abstract

The clinical phenotype in osteogenesis imperfecta (OI) is attributed to the dominant negative function of mutant type I collagen molecules in the extracellular matrix, by altering its structure and function. Intracellular retention of mutant collagen has also been reported, but its effect on cellular homeostasis is less characterized. Using OI patient fibroblasts carrying mutations in the α1(I) and α2(I) chains we demonstrate that retained collagen molecules are responsible for endoplasmic reticulum (ER) enlargement and activation of the unfolded protein response (UPR) mainly through the eukaryotic translation initiation factor 2 alpha kinase 3 (PERK) branch. Cells carrying α1(I) mutations upregulate autophagy, while cells with α2(I) mutations only occasionally activate the autodegradative response. Despite the autophagy activation to face stress conditions, apoptosis occurs in all mutant fibroblasts. To reduce cellular stress, mutant fibroblasts were treated with the FDA-approved chemical chaperone 4-phenylbutyric acid. The drug rescues cell death by modulating UPR activation thanks to both its chaperone and histone deacetylase inhibitor abilities. As chaperone it increases general cellular protein secretion in all patients' cells as well as collagen secretion in cells with the most C-terminal mutation. As histone deacetylase inhibitor it enhances the expression of the autophagic gene Atg5 with a consequent stimulation of autophagy. These results demonstrate that the cellular response to ER stress can be a relevant target to ameliorate OI cell homeostasis.

The mechanism for differential effect of nelfinavir and indinavir on collagen metabolism in human skin fibroblasts

“…Procollagen is degraded mainly in lysosomes but some of collagen mutant chains can be degraded in proteasomes . Interestingly, UPR induction due to retention of mutated collagen type I chains in ER was reported . Therefore, there is an open question whether impairing of collagen secretion may contribute to NEL‐induced ER stress in fibroblasts.…”

Genetic Disorders of the Extracellular Matrix

“…For example, with collagen I mutations in OI clear evidence of UPR activation was demonstrated for a procollagen I C‐propeptide trimerization mutation in the Aga2/+ mouse with upregulation of BiP, Hsp47, and Gadd153 (Chop) resulting in osteoblast apoptosis (Lisse et al, ). Recent studies on patient collagen I helix mutations reported upregulation of several components of the UPR, including BiP, PDI, ATF4, phospho‐PERK and XBP1 splicing (Besio et al, ). However, the pattern of upregulation of these individual UPR components was different in the five patient mutations studied.…”

“…However, the pattern of upregulation of these individual UPR components was different in the five patient mutations studied. For example, BiP was upregulated in only three of five COL1A1 or COL1A2 mutations tested (Besio et al, ), highlighting that while a UPR can be activated by these mutations, there is apparent mutation specificity in the strength and nature of the response. In a mouse model of mild to moderate OI due to a Col1a2 missense mutation (α2(I) G610C) which disturbs the collagen triple helix, ER accumulation of the mutant misfolded collagen I results in upregulation of CHOP, eiF2α phosphorylation, and chaperones αβ crystalline and HSP47, and it was concluded this ER‐stress did not involve the canonical UPR pathway (Mirigian et al, ).…”

“…Improving mutant protein folding by upregulating endogenous chaperones or by administering chemical chaperones that can stabilize proteins in their native conformation and rescue mutant protein folding and/or trafficking defects has been a widely used strategy in protein folding disorders (Perlmutter, ; Ulloa‐Aguirre et al, ; Arakawa et al, ; Rivas et al, ). The FDA‐approved chemical chaperone sodium 4‐phenylbutyrate (PBA) can reduce ER stress in vitro caused by COL1A1 OI mutations (Besio et al, ), COL4A2 (Murray et al, ), and COL4A5 (Wang et al, ) mutations. In a zebrafish model of OI, PBA administration ameliorated the skeletal pathology (Gioia et al, ).…”

Exogenous supply of Hsp47 triggers fibrillar collagen deposition in skin cell cultures in vitro

“…TGFbeta [16,17]) and chemical stimulants (e.g. ascorbic acid [17][18][19], glycolic acid [20], 4-phenyl butyric acid (4-PBA) [21] and retinol [22]) to boost the collagen production and matrix deposition. However, these molecules have multiple other roles in the body and the therapies are associated with negative side effects, such as promoting abnormal angiogenesis, or inflammatory responses.…”