REGULAR ARTICLE HDAC6 deacetylates alpha tubulin in sperm and modulates sperm motility in Holtzman rat Sweta Parab & Omshree Shetty & Reshma Gaonkar & Nafisa Balasinor & Vrinda Khole & Priyanka Parte Received: 10 July 2014 /Accepted: 15 October 2014 # Springer-Verlag Berlin Heidelberg 2014 Abstract Histone deacetylase 6 (HDAC6) is an alpha (α)- tubulin deacetylase and its over-expression has been demon- strated to promote chemotactic cell movement. Motility in sperm is driven by the flagella, the cytoskeletal structure comprising the microtubules, which are heterodimers of α- and β-tubulins. We have hypothesized that HDAC6, by virtue of being an α-tubulin deacetylase, might modulate sperm motility. However, the presence of HDAC6 on sperm has hitherto not been reported. In this study, we have demonstrat- ed, for the first time, the presence of HDAC6 transcript and protein in the testicular and caudal sperm of rat. We have observed a significantly overlapping expression of HDAC6 with acetyl α-tubulin (Ac α-tubulin) in the mid-piece and principal piece of sperm flagella, and the co-precipitation of α-tubulin and Ac α-tubulin together with HDAC6 and vice versa in sperm lysates. This indicates that HDAC6 interacts with α-tubulin. The HDAC6 activity of sperm, sperm motility and status of Ac α-tubulin investigated in the presence of HDAC inhibitors Trichostatin A, Tubastatin A and sodium butyrate demonstrate that HDAC6 in sperm is catalytically active and that inhibitors of HDAC6 increase acetylation and restrict sperm motility. Thus, we show that (1) active HDAC6 enzyme is present in sperm, (2) HDAC6 in sperm is able to deacetylate α-tubulin, (3) inhibition of HDAC6 results in increased Ac α-tubulin expression and (4) HDAC6 inhibition affects sperm motility. This evidence suggests that HDAC6 is involved in modulating sperm movement. Keywords Acetylated alpha-tubulin . Deacetylase activity . HDAC6 . HDAC inhibitor . Sperm motility . Rat Introduction Sperm flagellar motility is well-orchestrated and is attrib- utable to a highly organized microtubule-based structure called the axoneme. The axoneme is composed of 9 doublet microtubules and 2 singlet microtubules running along the length of the flagellum. The axonemal structure is surrounded by auxiliary dense fibres and the fibrous sheath that have no clear active role in the sliding of micro- tubules and flagellar movement. The microtubules are com- posed of α- and β-tubulins, which undergo several post- translational modifications, namely, polyglutamylation, polyglycylation, tyrosylation/detyrosylation and acetyla- tion/deacetylation. Whereas the polyglutamylation of the lateral chain of α-tubulin has been shown to have a role in flagellar motility, detyrosination and acetylation are thought to be important for the assembly of the axoneme (Gagnon et al. 1996). The distribution of acetylated (Ac) α- tubulin is tightly controlled and stereotypic. Ac α-tubulin is most abundant in stable microtubules but is absent from dynamic cellular structures (e.g. neuronal growth cones, leading edges of fibroblasts). Reversible acetylation of α- This work (RA/13/08-2013) was supported by grants from the Department of Science and Technology, India and the Indian Council of Medical Research. Junior Research Fellowship and Senior Research Fellowship provided to Sweta Parab and research associate fellowship to Dr. Omshree Shetty by the Department of Science and Technology is gratefully acknowledged. Electronic supplementary material The online version of this article (doi:10.1007/s00441-014-2039-x) contains supplementary material, which is available to authorized users. S. Parab : O. Shetty : V. Khole : P. Parte (*) Department of Gamete Immunobiology, National Institute for Research in Reproductive Health (ICMR), Mumbai 400012, India e-mail: partep@nirrh.res.in R. Gaonkar : N. Balasinor Department of Neuroendocrinology and Confocal Microscopy Lab, National Institute for Research in Reproductive Health (ICMR), Mumbai 400012, India Cell Tissue Res DOI 10.1007/s00441-014-2039-x