FIG 2.

Phylogenetic analysis places choanoflagellate NF-κBs as an outgroup of vertebrate and fly NF-κBs. (A) Bayesian analysis of holozoan RHDs, including the recently identified choanoflagellate NF-κBs. MEME analysis was performed on each sequence to identify shared motifs. NF-κB proteins and Rel proteins cluster separately from each other, and choanoflagellate NF-κBs cluster as a single outgroup. (B) A multiple-sequence alignment of important NF-κB subdomains across multicellular metazoans and single-celled premetazoans. Exact amino acid matches to either p100 or p105 are highlighted in blue across phyla, and residues with conserved functional groups are highlighted in yellow. All multicellular metazoans contain a defined start to the Rel homology domain (RHD), dimerization sequence, and nuclear localization sequence (NLS). Single-celled premetazoans have a less defined RHD start, low homology in the dimerization sequence, and no canonical NLS (with the exception of Capsaspora). All metazoan and premetazoan NF-κB sequences contain a highly conserved DNA-binding sequence.