TY - JOUR

T1 - A rare variant in human fibroblast activation protein associated with ER stress, loss of enzymatic function and loss of cell surface localisation

AU - Osborne, Brenna

AU - Yao, Tsun Wen

AU - Wang, Xin Maggie

AU - Chen, Yiqian

AU - Kotan, L. Damla

AU - Nadvi, Naveed A.

AU - Herdem, Mustafa

AU - McCaughan, Geoffrey W.

AU - Allen, John D.

AU - Yu, Denise M.T.

AU - Topaloglu, A. Kemal

AU - Gorrell, Mark D.

N1 - Funding Information: TWY and NAN were supported by Australian Postgraduate Awards. MDG was supported by project grants 512282 and 632822 from the Australian National Health and Medical Research Council (NHMRC) and grants from the Rebecca L. Cooper Medical Research Foundation . GWM is supported by NHMRC program grant 571408 . We thank Professor John Bateman, Murdoch Children's Research Institute, Melbourne, Australia for advice. Dr Ben Roediger and Dr Volker Benseler of Centenary Institute, and Eleanor Kable of Australian Centre for Microscopy & Microanalysis, University of Sydney, assisted with confocal image analysis.

PY - 2014/7

Y1 - 2014/7

N2 - Fibroblast activation protein (FAP) is a focus of interest as a potential cancer therapy target. This membrane bound protease possesses the unique catalytic activity of hydrolysis of the post-proline bond two or more residues from the N-terminus of substrates. FAP is highly expressed in activated fibroblastic cells in tumours, arthritis and fibrosis. A rare, novel, human polymorphism, C1088T, encoding Ser363 to Leu, occurring in the sixth blade of the β propeller domain, was identified in a family. Both in primary human fibroblasts and in Ser363LeuFAP transfected cells, we showed that this single substitution ablates FAP dimerisation and causes loss of enzyme activity. Ser363LeuFAP was detectable only in endoplasmic reticulum (ER), in contrast to the distribution of wild-type FAP on the cell surface. The variant FAP showed decreased conformational antibody binding, consistent with an altered tertiary structure. Ser363LeuFAP expression was associated with upregulation of the ER chaperone BiP/GRP78, ER stress sensor ATF6, and the ER stress response target phospho-eIF2α, all indicators of ER stress. Proteasomal inhibition resulted in accumulation of Ser363LeuFAP, indicating the involvement of ER associated degradation (ERAD). Neither CHOP expression nor apoptosis was elevated, so ERAD is probably important for protecting Ser363LeuFAP expressing cells. These data on the first loss of function human FAP gene variant indicates that although the protein is vulnerable to an amino acid substitution in the β-propeller domain, inactive, unfolded FAP can be tolerated by cells.

AB - Fibroblast activation protein (FAP) is a focus of interest as a potential cancer therapy target. This membrane bound protease possesses the unique catalytic activity of hydrolysis of the post-proline bond two or more residues from the N-terminus of substrates. FAP is highly expressed in activated fibroblastic cells in tumours, arthritis and fibrosis. A rare, novel, human polymorphism, C1088T, encoding Ser363 to Leu, occurring in the sixth blade of the β propeller domain, was identified in a family. Both in primary human fibroblasts and in Ser363LeuFAP transfected cells, we showed that this single substitution ablates FAP dimerisation and causes loss of enzyme activity. Ser363LeuFAP was detectable only in endoplasmic reticulum (ER), in contrast to the distribution of wild-type FAP on the cell surface. The variant FAP showed decreased conformational antibody binding, consistent with an altered tertiary structure. Ser363LeuFAP expression was associated with upregulation of the ER chaperone BiP/GRP78, ER stress sensor ATF6, and the ER stress response target phospho-eIF2α, all indicators of ER stress. Proteasomal inhibition resulted in accumulation of Ser363LeuFAP, indicating the involvement of ER associated degradation (ERAD). Neither CHOP expression nor apoptosis was elevated, so ERAD is probably important for protecting Ser363LeuFAP expressing cells. These data on the first loss of function human FAP gene variant indicates that although the protein is vulnerable to an amino acid substitution in the β-propeller domain, inactive, unfolded FAP can be tolerated by cells.

KW - Dipeptidyl peptidase

KW - Endoplasmic reticulum associated degradation

KW - Endoplasmic reticulum stress

KW - Fibroblast activation protein

KW - Polymorphism

KW - Unfolded protein response

UR - http://www.scopus.com/inward/record.url?scp=84899809867&partnerID=8YFLogxK

U2 - 10.1016/j.bbapap.2014.03.015

DO - 10.1016/j.bbapap.2014.03.015

M3 - Journal article

C2 - 24717288

AN - SCOPUS:84899809867

VL - 1844

SP - 1248

EP - 1259

JO - B B A - Proteins and Proteomics

JF - B B A - Proteins and Proteomics

SN - 1570-9639

IS - 7

ER -