Comparison of NF-κB from the protists Capsaspora owczarzaki and Acanthoeca spectabilis reveals extensive evolutionary diversification of this transcription factor
Authors: Williams Leah M. , Sridhar Sainetra , Samaroo Jason , Peart Jada , Adindu Ebubechi K. , Addanki Anvitha , DiRusso Christopher J. ,   BB522 Molecular Biology Laboratory, Alburi Dana H. M. , Anisimov Ludmila , Armstrong Aria Y. , Badger Sydney J. , Banaie Elham , Teixeira Joana A. Barbosa , Billingsley Madeleine T. , Calikyan Anoush , Chen Yinxing , Coia Aidan B. , Cutillo Daniel , Dooling Breanna R. , Doshi Parth P. , Dubinsky Kyra R. , Velasco Berta Escude , Evans Jabari R. , Gordon Jasmine , Guan Huibo , Haliotis Spiro N. , Hood Niccolas T. , Huang Yen-Chun , Jiang Wenjing , Kreber Isabelle C. , Kurak Ekin B. , Lee Cheng-Che , Lehmann Tanner M. , Lin Savina J. W. , Liu Edward , Liu Kevin , Liu Yen-Yu , Luther Alexandra L. , Macgranaky-Quaye Alexa A. , Magat Daniel J. , Malsick Lauren E. , Masoudi Parmida , Masoudi Parsida , Max Chad R. H. , McCaslin Ethan Z. , McGeary Eleanor T. , McLaughlin Kathleen M. , Momyer Victoria S. A. , Murphy Lake D. , Nguyen Sonny V. , Ni Kareemah , Novak Leon , Santos Roberto Nunes Campos E. , Osayame Yemi D. , Chang Jun Bai Park , Patel Harshal M. , Pham Tony V. , Phillips Sheila M. , Piedrahita Jhonathan Perea , Post Tricia L. , Prather Rebecca A. , Reck Pauline I. , Rodriguez Jaime A. , Rolle Kirquenique A. , Salzo Joseph A. , Satko Kathryn M. , Settipane Davis G. , Sevola Kara J. , Shah Mithil V. , Skidanova Viktoriya , Snyder Georgia M. , Sprague Rebecca J. , Stagg Ryan A. , Tong Danielle , Towers Andreas A. , Turgiss Nicholas W. , Wheeler Natalie S. , Yung Ann S. , Aguirre Carrión Pablo J. , Rodriguez-Sastre Nahomie , Siggers Trevor , Gilmore Thomas D. 
Published: December 2021

We provide a functional characterization of transcription factor NF-κB in protists and provide information about the evolution and diversification of this biologically important protein. We characterized NF-κB in two protists using phylogenetic, cellular, and biochemical techniques. NF-κB of the holozoan Capsaspora owczarzaki (Co) has an N-terminal DNA-binding domain and a C-terminal Ankyrin repeat (ANK) domain, and its DNA-binding specificity is more similar to metazoan NF-κB proteins than to Rel proteins. Removal of the ANK domain allows Co-NF-κB to enter the nucleus, bind DNA, and activate transcription. However, C-terminal processing of Co-NF-κB is not induced by IκB kinases in human cells. Overexpressed Co-NF-κB localizes to the cytoplasm in Co cells. Co-NF-κB mRNA and DNA-binding levels differ across three Capsaspora life stages. RNA-sequencing and GO analyses identify possible gene targets of Co-NF-κB. Three NF-κB-like proteins from the choanoflagellate Acanthoeca spectabilis (As) contain conserved Rel Homology domain sequences, but lack C-terminal ANK repeats. All three As-NF-κB proteins constitutively enter the nucleus of cells, but differ in their DNA-binding abilities, transcriptional activation activities, and dimerization properties. These results provide a basis for understanding the evolutionary origins of this key transcription factor and could have implications for the origins of regulated immunity in higher taxa.