Disruption of Ttll5/Stamp gene (Tubulin tyrosine ligase-like protein 5/SRC-1 and TIF2 associated modulatory protein gene) in male mice causes sperm malformation and infertility

Tubulin tyrosine ligase-like family member 5 (TTLL5/STAMP) has multiple activities in cells. TTLL5 is one of 13 TTLLs, has polyglutamylation activity, augments the activity of p160 coactivators (SRC-1 and TIF2) in glucocorticoid receptor-regulated gene induction and repression, and displays steroid-independent growth activity with several cell types. To examine TTLL5/STAMP functions in whole animals, mice were prepared with an internal deletion that eliminated several activities of the Stamp gene. This mutation causes both reduced levels of STAMP mRNA and C-terminal truncation of STAMP protein. Homozygous targeted mutant (Stamptm/tm) mice appear normal except for marked decreases in male fertility associated with defects in progressive sperm motility. Abnormal axonemal structures with loss of tubulin doublets occur in most Stamptm/tm sperm tails in conjunction with substantial reduction in tubulin polyglutamylation, which closely correlates with the reduction in mutant STAMP mRNA. The axonemes in other structures appear unaffected. There is no obvious change in the organs for sperm development of wt vs. Stamptm/tm males despite the levels of wt STAMP mRNA in testes being 20-fold higher than in any other organ examined. This defect in male fertility is unrelated to other Ttll genes or 24 genes previously identified as important for sperm function. Thus, STAMP appears to participate in a unique, tissue-selective TTLL-mediated pathway for tubulin polyglutamylation that is required for sperm maturation and motility and may be relevant for male fertility. Testes RNA samples for EXON arrays from WT and KO with three biological replications微管酪氨酸连接酶样家族成员5（Tubulin tyrosine ligase-like family member 5，TTLL5/STAMP）在细胞内具备多种生物学功能。TTLL5作为13种TTLL家族蛋白之一，不仅具有多谷氨酰化修饰活性，还可增强p160辅激活因子（p160 coactivators，SRC-1与TIF2）在糖皮质激素受体（glucocorticoid receptor）调控的基因诱导与抑制过程中的活性，同时在多种细胞类型中表现出类固醇非依赖性生长活性。为探究TTLL5/STAMP在完整生物体中的功能，研究人员制备了携带内部缺失突变的小鼠，该突变消除了Stamp基因的多种活性。此突变可同时降低STAMP mRNA的表达水平，并导致STAMP蛋白发生C端截短。纯合靶向突变（Stamptm/tm）小鼠除雄性生育能力显著下降（伴随精子渐进式运动能力缺陷）外，其余表型均无异常。在大多数Stamptm/tm小鼠的精子尾部中，可观察到异常轴丝结构，表现为微管双联微管缺失，同时伴随微管多谷氨酰化修饰水平的显著降低，该现象与突变型STAMP mRNA的表达水平降低高度相关。而其他组织中的轴丝结构未受明显影响。尽管野生型（wild type，wt）小鼠睾丸中的wt STAMP mRNA表达水平较其他所有检测器官高出20倍，但野生型与Stamptm/tm雄性小鼠的精子发育器官并无明显异常。该雄性生育缺陷与其他Ttll家族基因以及此前已报道的24个精子功能相关基因均无关联。综上，STAMP似乎参与了一条独特的、组织选择性的TTLL介导的微管多谷氨酰化修饰通路，该通路对于精子成熟与运动至关重要，可能与雄性生育能力密切相关。本研究用于外显子阵列（EXON arrays）检测的睾丸RNA样本均来自野生型与敲除（Knock Out，KO）小鼠，且包含3次生物学重复。