Aberrant Expression of Dynein light chain 1 (DYNLT1) is Associated with Human Male Factor Infertility*

DYNLT1 is a member of a gene family identified within the t-complex of the mouse, which has been linked with male germ cell development and function in the mouse and the fly. Though defects in the expression of this gene are associated with male sterility in both these models, there has been no study examining its association with spermatogenic defects in human males. In this study, we evaluated the levels of DYNLT1 and its expression product in the germ cells of fertile human males and males suffering from spermatogenic defects. We screened fertile (n = 14), asthenozoospermic (n = 15), oligozoospermic (n = 20) and teratozoospermic (n = 23) males using PCR and Western blot analysis. Semiquantitative PCR indicated either undetectable or significantly lower levels of expression of DYNLT1 in the germ cells from several patients from across the three infertility syndrome groups, when compared with that of fertile controls. DYNLT1 was localized on head, mid-piece, and tail segments of spermatozoa from fertile males. Spermatozoa from infertile males presented either a total absence of DYNLT1 or its absence in the tail region. Majority of the infertile individuals showed negligible levels of localization of DYNLT1 on the spermatozoa. Overexpression of DYNLT1 in GC1-spg cell line resulted in the up-regulation of several cytoskeletal proteins and molecular chaperones involved in cell cycle regulation. Defective expression of DYNLT1 was associated with male factor infertility syndromes in our study population. Proteome level changes in GC1-spg cells overexpressing DYNLT1 were suggestive of its possible function in germ cell development. We have discussed the implications of these observations in the light of the known functions of DYNLT1, which included protein trafficking, membrane vesiculation, cell cycle regulation, and stem cell differentiation.The t-complex of the mouse occupies the proximal half of chromosome 17 and contains genes which have profound effects on spermatogenesis. Multiple mutations in several loci in the t-complex appear to interact to cause complete male sterility (1, 2). Tctex-1 (t-complex testis expressed-1), lately renamed as dynein light chain 1 (Dynlt1)¹, is identified as a candidate gene involved in male sterility in mice (1) and maps to the t-complex in mice (3). Dynlt1 is a member of a multigene family which is virtually germ cell-specific and is eightfold over expressed in t-homozygotes and 200-fold higher in testis than in other adult tissues (1). The human homologue of the mouse Dynlt1 is located on chromosome 6q25.2–25.3. The amino acid sequence shows a high degree of similarity to the predicted product of the Dynlt1 gene of the mouse t complex (4).DYNLT1 gene encodes a 14 kDa protein constituting the inner arm L1 of cytoplasmic and flagellar dynein complexes (5, 6). DYNLT1 is localized to Golgi complexes as well (7). DYNLT1 protein is present in sperm tails and oocytes (8, 9). A wide range of cellular events are brought about by cytoplasmic dynein and its association with the accessory intermediate, light intermediate, and light chain subunits. These subunits define the interaction of cytoplasmic dynein motor complex with other molecules (10). DYNLT1 is involved in cargo binding (11), lymphocyte division (8), vesicle transport (12–14), and human embryo implantation (15). DYNLT1 is known to undergo phosphorylation during apical delivery of rhodopsin (16) and during its interaction with the bone morphogenetic receptor type II (BMPRII) (17). DYNLT1 can function in dynein-independent fashion as a cell fate regulator by its interaction with G-protein β γ subunit regulating initial neurite sprouting (18), axonal specification, and elongation of hippocampal neurons in culture (11, 19). GEF-H1 is bound to microtubules by DYNLT1 and its release without microtubule depolymerization is mediated through the interaction of DYNLT1 with G proteins (20). DYNLT1 is a novel marker for neural progenitors in adult brain (21). DYNLT1 regulatory element was identified which selectively marked nestin⁺/GFAP⁺/Sox2⁺ neural stem-like cells in developing and adult brain (22). The genetic knockdown of DYNLT1 in radial precursors promoted neurogenesis (23). The use of GFP placed under the control of DYNLT1 promoter to mark adult neural stem cells and thus allowing the insertion of any nucleotide sequence selectively into neural progenitors has been patented (24).DYNLT1 is reported to have functional roles in non-murine germ cells as well. DYNLT1 was found to be essential during spermatid differentiation in Drosophila (10) and a mouse DYNLT1 homolog was identified in the dynein light chain of sea urchin sperm flagella (25, 26). However, the expression of DYNLT1 in human testicular germ cells and its association, if any, with human male factor subfertility are not yet evaluated. This study evaluates the association between DYNLT1expression and spermatogenesis in infertile human males and the possible function of DYNLT1 in spermatogonial cell division and differentiation.