Gelsolin domain 2 Ca2+ affinity determines susceptibility to furin proteolysis and familial amyloidosis of finnish type.
Mutation of aspartic acid 187 to asparagine (D187N) or tyrosine (D187Y) in domain 2 of the actin-modulating protein gelsolin causes the neurodegenerative disease familial amyloidosis of Finnish type (FAF). These mutations render plasma gelsolin susceptible to aberrant proteolysis by furin in the trans-Golgi network, the initial proteolytic event in the formation of 71 and 53 residue fragments that assemble into amyloid fibrils. Ca(2+) binding stabilizes wild-type domain 2 gelsolin against denaturation and proteolysis, but the FAF variants are unable to bind and be stabilized by Ca(2+). Though the chain of events initiating FAF has been elucidated recently, uncertainty remains about the mechanistic details that allow the FAF variants to be processed. To test the hypothesis that impaired Ca(2+) binding in the D187 variants, but not other factors specific to residue 187, increases susceptibility to aberrant proteolysis and subsequent amyloidogenesis, we designed the gelsolin variant E209Q to remove a different Ca(2+) ligand from the same Ca(2+) site that is affected in the FAF variants. Here, we show that E209Q domain 2 does not bind Ca(2+) and is not stabilized against denaturation or furin proteolysis, analogous to the behavior exhibited by the FAF variants. Transfection of full-length E209Q into COS cells results in secretion of both the full-length and furin-processed fragments, as observed with D187N and D187Y. Mutation of the furin consensus sequence in D187N and E209Q gelsolin prevents cleavage during secretion, indicating that inhibition of proprotein convertases (furin) represents a viable therapeutic approach for the treatment of FAF. Mutations that diminish domain 2 Ca(2+) binding allow furin access to an otherwise protected cleavage site, initiating the proteolytic cascade that leads to gelsolin amyloidogenesis and FAF.
Department of Chemistry and Skaggs Institute for Chemical Biology, The Scripps Research Institute BCC265, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Mary E
ME
Huff
Lesley J
LJ
Page
William E
WE
Balch
Jeffery W
JW
Kelly
D028227:Amyloid Neuropathies, Familial / Q000378:metabolism*; D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000378:metabolism*; D016384:Consensus Sequence; D045683:Furin / Q000378:metabolism*; D018260:Gelsolin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary
D018260:Gelsolin; D045683:Furin; D002118:Calcium