THE TESTIS-SPECIFIC HIGH-MOBILITY-GROUP PROTEIN, A PHOSPHORYLATION-DEPENDENT DNA-PACKAGING FACTOR OF ELONGATING AND CONDENSING SPERMATIDS

Mammalian spermiogenesis is characterized by a striking restructuring of the spermatid chromatin caused by the replacement of nucleohistones with transition proteins and their subsequent replacement with nucleo -protamines. The onset of nuclear elongation and chromatin condensatio n in spermatids is accompanied by a general decrease in the transcript ional activity of the DNA. A recently identified testis-specific high- mobility-group (tsHMG) protein, similar to the human mitochondrial tra nscription factor I and to the linker-associated protein delta of Tetr ahymena thermophila micronuclei, is thought to play a structural role in this process. We confirm by immunoblot analysis of fractionated ger m cells that the presence of tsHMG is restricted to transcriptionally quiescent elongating and condensing spermatids. Purified recombinant t sHMG protein displays preferential binding to supercoiled plasmid DNA, which reversibly protects the DNA against the DNA-reIaxing activity o f eukaryotic topoisomerase I and also impairs the transcriptional acti vity of this template when assayed in vitro. The tsHMG protein can als o introduce negative supercoils into a relaxed plasmid substrate in a topoisomerase I-dependent manner. We also show that the tsHMG protein is the substrate of a Ca2+-phospholipid-dependent protein kinase (prot ein kinase C) present in testis extracts of adult mice and demonstrate that phosphorylation by protein kinase C is required for both the DNA -binding and the topoisomerase I-dependent supercoiling activities of tsHMG. Our results support the hypothesis that the spermatid tsHMG pro tein is a topological factor (transition protein) that can modulate th e activity of topoisomerase I. This activity could contribute to the i mportant transition in chromatin structure which leads to the decrease in DNA metabolism observed at the early stages of spermatid elongatio n.