Investigating The Regulation Of Meiosis By A Novel Synaptonemal Complex Component HTP-4

Meiosis is a specialized cell division process for the production of sexual reproduction.Errors during meiotic prophase could affect the accuracy of chromosome segregation,leading to the production of aneuploid gametes,which may further result in serious reproductive health problems such as miscarriages and birth defects.Therefore,the study of meiosis is of great significance to human reproduction.C.elegans is a mature model organism in the field of meiosis research.The germ cells in the gonads exhibit a temporal/spatial gradient arrangement,and all stages of meiotic prophase can be observed in a single gonad.In this study,by using high specific immunoprecipitation and mass spectrometry analysis,we identified a new component of the synaptonemal complex named HTP-4,which has a conserved HORMA domain.In order to investigate the function of htp-4 gene and the expression pattern of HTP-4,CRISPR gene editing technology was used to create htp-4 mutant strain and htp-4::gfp transgenic strain.We found that HTP-4 is not expressed in the embryos and intestines,but only in the gonads,indicating that HTP-4 is a germline-specific protein.In the gonads,HTP-4expression started to be observed at leptotene/zygotene,and enhanced significantly during late meiotic prophase,exhibiting a dynamic change in its expression level.HTP-4 exhibites parrellel tracks on pachytene chromosomes and colocalizes with HTP-3,confirming that HTP-4 is a lateral element of the synaptonemal complex.DAPI staining of htp-4 mutant goands revealed a shortened transition zone,suggesting that HTP-4may function as a synapsis checkpoint protein during early meiotic prophase.In mutants that are deficient in synapsis,deletion of HTP-4 also results in shortened transition zone,further supports our hypothesis that HTP-4 acts as a checkpoint protein.In addition,we found that the synaptonemal complex can assemble normally in htp-4mutants,indicating that homologous synapsis is independent of HTP-4.In the htp-4mutants,six COSA-1::GFP foci can also be observed in pachytene nuclei,and six DAPI-stained bodies were also observed in diakinesis nuclei,indicating that HTP-4 is not required for the formation of crossovers.Microscopy analysis revealed that HTP-4::GFP undergoes significant changes in its chromosomal localization during meiosis.From the transition zone to the end of the pachytene,HTP-4 localizes to the whole chromosome axis;while during late prophase where chromosomes undergo remodeling process,HTP-4 gradually localizes to the long arms of the bivalents.Interestingly,in syp-1 mutants that lack the synaptonemal complex,HTP-4 cannot localize to the chromosome axis properly before pachytene stage.However,it can still be properly recruired to the chromosomes after diplotene stage.These and other results suggested that the localization of HTP-4 during early meiotic prophase depends on the assembly of the synaptonemal complex and it may function as a checkpoint protein sensing the chromosome synapsis status.While during late meiotic prophase,HTP-4 reaches its maximal expression and localizes to the long arms of the bivalents,where it may act as a structural component of the chromosome axis and be involved in chromosome remodeling process.These findings broaden our knowledge about mechanisms controlling homologous chromosome synapsis and provide novel insights about genome integrity maintenance during meiosis.