Title : A Multiprotein Complex Regulates Interference-sensitive Crossover Formation 6 in Rice

36 In most eukaryotes, a set of conserved proteins that are collectively termed ZMM 37 proteins (named after ZIP1/2/3/4, MSH4/5, MER3, and SPO16 in Saccharomyces 38 cerevisiae) are essential for the formation of the majority of meiotic crossovers (COs). 39 Recent reports indicated that ZIP2 acts together with SPO16 and ZIP4 to control CO 40 formation through recognizing and stabilizing early recombination intermediates in 41 budding yeast. However, whether this mechanism is conserved in plants is not clear. 42 Here, we characterized the functions of OsSHOC1 (ZIP2 ortholog), OsPTD (SPO16 43 ortholog) and their interactions with other ZMM proteins in rice (Oryza sativa). We 44 demonstrated that disruption of OsSHOC1 caused a reduction of CO numbers to ~ 83% 45 of wild type, whereas synapsis and early meiotic recombination steps were not affected. 46 Furthermore, OsSHOC1 interacts with OsPTD, which is responsible for the same set of 47 CO formations as OsSHOC1. Additionally, OsSHOC1 and OsPTD are required for the 48 normal loading of other ZMM proteins, and conversely, the localizations of OsSHOC1 49 and OsPTD were also affected by the absence of OsZIP4 and OsHEI10. OsSHOC1 50 interacts with OsZIP4, OsMSH5 ,and PTD interacts with OsHEI10. Furthermore, BiFC 51 and yeast-three hybrid assays demonstrated that OsSHOC1, OsPTD, OsHEI10, and 52 OsZIP4 were able to form various combinations of heterotrimers. Moreover, statistical 53 and genetic analysis indicated that osshoc1-2 and osptd-1 are epistatic to oshei10 and 54 www.plantphysiol.org on July 4, 2019 Published by Downloaded from Copyright © 2019 American Society of Plant Biologists. All rights reserved. 3 oszip4 in meiotic CO formation. Taken together, we propose that OsSHOC1, OsPTD, 55 OsHEI10, and OsZIP4 form multiple protein complexes that have conserved functions 56 in promoting class I CO formation. 57 58