Speedy A governs non-homologous XY desynapsis as a unique prerequisite for XY loop-axis organization

In mouse early pachytene spermatocytes, the X and Y chromosomes undergo rapid non-homologous (NH) synapsis and desynapsis, but the functional significance remains unknown. Here, we report that pachynema-specific knockout of Speedy A (SpdyA) from telomeres caused persistent Y-X NH synapsis, with the entire Y axis synapsed onto the X axis. This persistent Y-X NH synapsis did not interrupt meiotic sex chromosome inactivation, recombination, or sex body formation, but it disrupted X-Y loop-axis organization and homologous X-Y desynapsis, leading to spermatocyte death. Similarly, persistent Y-X NH synapsis was also observed in pachytene spermatocytes lacking TRF1, where SpdyA was frequently lost from X-Y non-pseudoautosomal region (non-PAR) telomeres. Mechanistic studies revealed that the Serine 48 of SUN1 is a key SpdyA/CDK2 phosphorylation site needed for Y-X NH desynapsis. We propose that SpdyA governs Y-X NH desynapsis by stabilizing the linkage between the X-Y non-PAR telomeres and their LINC complexes, and that this process is regulated independently from other aspects of pachynema progression. Our findings suggest a key role for Y-X NH desynapsis in establishing proper X-Y loop-axis organization. Overall design: To investigate whether meiotic sex chromosome inactivation (MSCI) is disrupted by persistent full Y-X non-homologous (NH) synapsis following Spdya deletion, synchronized Spdyafl/fl (control) and SpdyacKO testes were collected on RA11.5 (i.e., 11.5 days post RA restoration, equivalent to mid-late pachytene stage) and subjected to RNA-Seq analysis. Four mice were used for each group.