© The Author(s) 2022. Published by Oxford University Press on behalf of National Academy of Sciences. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. ORIGINAL UNEDITED MANUSCRIPT Main Manuscript for Beyond a platform protein for the degradosome assembly: The Apoptosis Inducing Factor as efficient nuclease involved in chromatinolysis Nerea Novo a,b , Silvia Romero-Tamayo a,b , Carlos Marcuello c,d , Sergio Boneta a , Irene Blasco- Machin a , Adrián Velázquez-Campoy a,b,e,f , Raquel Villanueva a,b , Raquel Moreno-Loshuertos a , Anabel Lostao c,d,g , Milagros Medina a,b* and Patricia Ferreira a,b* . a Departamento de Bioquímica y Biología Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, Spain. b Instituto de Biocomputación y Física de Sistemas Complejos, BIFI (GBsC-CSIC Joint Unit), Universidad de Zaragoza, Spain. c Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, 5009 Zaragoza, Spain d Laboratorio de Microscopias Avanzadas (LMA), Universidad de Zaragoza, 50018 Zaragoza, Spain e Aragón Institute for Health Research (IIS Aragón), Zaragoza, Spain. f Biomedical Research Networking Centre for Liver and Digestive Diseases (CIBERehd), Madrid, Spain g Fundación ARAID, Aragón, Spain *Milagros Medina and *Patricia Ferreira. Email: mmedina@unizar.es and ferreira@unizar.es Author Contributions: MM and PF designed research; NN, SRT, CM, SB, IBM, RV, RML performed research; NN, SRT, CM, AVC, AL, MM and PF analyzed data; NN, CM, AL, MM and PF wrote the original paper; NN, CM, AL, AVC, RV, RML, MM and PF review the paper. Competing Interest Statement: The authors declare no competing interest. Classification: Biological Sciences, Biochemistry. Keywords: human Apoptosis Inducing Factor, DNA-degradosome complex, nuclease activity, chromatinolysis. This word file includes: Main Text Figure Legends 1 to 4 Abstract The Apoptosis-Inducing Factor (AIF) is a moonlighting flavoenzyme involved in the assembly of mitochondrial respiratory complexes in healthy cells, but also able to trigger DNA cleavage and parthanatos. Upon apoptotic-stimuli, AIF redistributes from the mitochondria to the nucleus, where upon association with other proteins such as endonuclease CypA and histone H2AX, it is proposed to organize a DNA-degradosome complex. In this work, we provide evidence for the molecular assembly of this complex as well as for the cooperative effects among its protein components to degrade genomic DNA into large fragments. We have also uncovered that AIF has nuclease activity that is stimulated in the presence of either Mg 2+ or Ca 2+ . Such activity allows Downloaded from https://academic.oup.com/pnasnexus/advance-article/doi/10.1093/pnasnexus/pgac312/6961068 by guest on 28 December 2022