title: Interspecies Variation in the Functional Consequences of Mutation of Cytochrome c.
abstract: The naturally occurring human cytochrome c variant (G41S) is associated with a mild autosomal dominant thrombocytopenia (Thrombocytopenia Cargeeg) caused by dysregulation of platelet production. The molecular basis of the platelet production defect is unknown. Despite high conservation of cytochrome c between human and mouse (91.4% identity), introducing the G41S mutation into mouse cytochrome c in a knockin mouse (CycsG41S/G41S) did not recapitulate the low platelet phenotype of Thrombocytopenia Cargeeg. While investigating the cause of this disparity we found a lack of conservation of the functional impact of cytochrome c mutations on caspase activation across species. Mutation of cytochrome c at residue 41 has distinct effects on the ability of cytochrome c to activate caspases depending on the species of both the cytochrome c and its binding partner Apaf-1. In contrast to our previous results showing the G41S mutation increases the ability of human cytochrome c to activate caspases, here we find this activity is decreased in mouse G41S cytochrome c. Additionally unlike wildtype human cytochrome c, G41S cytochrome c is unable to activate caspases in Xenopus embryo extracts. Taken together these results demonstrate a previously unreported species-specific component to the interaction of cytochrome c with Apaf-1. This suggests that the electrostatic interaction between cytochrome c and Apaf-1 is not the sole determinant of binding, with additional factors controlling binding specificity and affinity. These results have important implications for studies of the effects of cytochrome c mutations on the intrinsic apoptosis pathway.
authors.affiliation: Department of Biochemistry, Otago School of Medical Sciences, University of Otago, Dunedin, New Zealand.
authors.forename: Tracy M
authors.initials: TM
authors.lastname: Josephs
authors.affiliation: Department of Biochemistry, Otago School of Medical Sciences, University of Otago, Dunedin, New Zealand.
authors.forename: Moira E
authors.initials: ME
authors.lastname: Hibbs
authors.affiliation: Department of Biochemistry, Otago School of Medical Sciences, University of Otago, Dunedin, New Zealand.
authors.forename: Lily
authors.initials: L
authors.lastname: Ong
authors.affiliation: Department of Pathology, Dunedin School of Medicine, University of Otago, Dunedin, New Zealand.
authors.forename: Ian M
authors.initials: IM
authors.lastname: Morison
authors.affiliation: Department of Biochemistry, Otago School of Medical Sciences, University of Otago, Dunedin, New Zealand.
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Ledgerwood
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D045304:Cytochromes c / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D055879:Gene Knock-In Techniques; D006410:Hematopoiesis; D006801:Humans; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009154:Mutation; D010976:Platelet Count; D013045:Species Specificity
chemical_list: D045304:Cytochromes c; D020169:Caspases