Fig. 5: Three of the four highest frequency missense human MLKL SNPs encode non-conservative amino acid substitutions within or adjacent to the brace helix region.
a S132 and R146 (magenta) are located on either side of D140 (yellow—equivalent to mouse D139) in the first human MLKL brace helix. Alternate amino acids encoded by human polymorphisms indicated in parentheses. b G202 is predicted to be on an α helix unique to MLKL2 isoform and to form an interface along with S132 and R146. The mouse equivalent of human rs35589326 (hMLKLS132P), mMLKLS131P, spontaneously forms membrane-associated high molecular weight complexes following Blue Native (BN) PAGE (c) and kills MDFs (d) in the absence of extrinsic necroptotic stimuli when expressed in mouse dermal fibroblasts for 6 (c) and 21 hrs respectively (d). C; cytoplasmic fraction, M; crude membrane fraction, TSI; TNF, Smac-mimetic and IDN6556, Chlor: Chloroquine. c Representative of two independent experiments with similar results. Error bars in (d) indicate the mean ± SEM of 4–5 independent experiments. e Schematic showing brace helix variant combinations identified as alleles in trans in three CRMO patients. f MTRs are mapped onto the structure of MLKL to show regions that have low tolerance to missense variation in the human population (red) and regions that have increased tolerance to missense variation (blue), normalized to the gene’s MTR distribution. g Multiple sequence alignment (MSA) conservation scores are mapped onto the structure of MLKL to show regions that are highly conserved through evolution (red) and regions that are less conserved through evolution (blue).
