Opeyemi Dhikhirullahi ¹; Ava Miciuda ¹; Cheng Zheng ^{1, 2}; Wei Li ¹; Zhibing Zhang ^{1, 3}

1. Department of Physiology, Wayne State University, Detroit, MI, 48201, USA;


2. School of Public Health, Wuhan University of Science and Technology, Wuhan, Hubei, China;


3. Department of Obstetrics & Gynecology, Wayne State University, Detroit, MI, 48201, USA.

The currently available contraceptives for men besides sex abstinence/withdrawal are condoms and vasectomy; however, these approaches have limitations in terms of invasiveness, consistency of usage, and dependability. This begs the need for the development of a safe, effective and reversible drug. A mature functional spermatozoon develops through numerous dynamic stages involving a number of genes that regulate the process. The genes that play roles in late stages of spermatogenesis are ideal targets for developing contraceptives because it is likely to result in fewer adverse effects and be reversible. Mouse Meiosis expressed gene 1 (MEIG1) is a small protein that is indispensable for spermiogenesis. A yeast two-hybrid screen using the full-length mouse MEIG1 as bait identified PACRG as a major binding partner. From the same yeast two-hybrid screen, an RNA splicing factor SC35 was also identified. The two proteins colocalize in the nuclei of spermatocytes and round spermatids, as shown by immunofluorescence staining. Interaction between MEIG1 and SC35 was further evaluated using a luciferase-based protein complementary assay. MEIG1 was fused to the N-fragment of luciferase (MEIG1-N-Luc), and SC35 was fused to the C-fraction of luciferase (SC35-C-Luc). The HEK293 cells transfected with each plasmid did not show luciferase activity. However, robust luciferase activity was recorded in the cells co-transfected with the two plasmids, indicating that SC35 and MEIG1 interact to bring N and C fractions of the luciferase to restore the full luciferase. Twelve amino acids on the MEIG1 protein surface are believed to mediate interactions with other proteins. These amino acids were mutated, and the interaction between SC35 with these mutant MEIG1 was examined. We discovered that some mutations significantly reduced interaction. We further confirmed that the interaction was conserved between human MEIG1 and SC35. Luciferase activity was also reconstituted in vitro using lysates from HEK cells transfected with individual plasmids expressing the two fusion proteins. Interaction between MEIG1 and SC35 indicates that MEIG1 may also be involved in the RNA splicing process required for the translation of crucial proteins that are required for sperm maturation. Targeting MEIG1-SC35 interaction might be another strategy for the development of male-based contraceptives.