N-ethyl-nitrosourea (ENU) mutagenesis was carried out in an effort to identify genes that are critical for male fertility. Our lab has successfully identified the Kaos mouse line that carries a C→T mutation in exon 14 of the Leucine rich repeats and guanylate kinase domain containing (Lrguk) gene. The mutation results in the introduction of a premature stop codon at amino acid position 528 on the LRGUK protein.
Kaos homozygous mutant males are sterile as determined by breeding experiments. Mutant males have reduced testis weights and 80% reduction in daily sperm production. Histological analysis showed that the sperm from Kaos mutant have severe sperm head and tail morphological abnormalities. Mutant sperm tail appeared coiled and had no capacity for motility. Sperm ultrastructure by EM revealed the mutant germ cells has a disorganized manchette and a constriction of the perinuclear ring and a complete absence of axoneme formation.
Lrguk mRNA is predominantly expressed in haploid germ cells in the testis but also expressed at lower levels in ciliated tissues including the lung, trachea and ovary. Immunofluorescence staining showed that LRGUK is localised in the sperm tail and manchette.
Further investigations showed that the Kaos homozygous males develop hydrocephalus. Magnatic Resonance Imaging and histological analysis revealed significantly enlargement of the brain ventricle and disorganized cilia and is consistent with a role for LRGUK in motile cilia formation more broadly.
In an effort to define the biochemical function of LRGUK-1 we have identified several binding partners using the yeast two-hybrid (Y2H) method. These include HOOK2 which has a critical role in basal body extension. Together, our data suggests a role of LRGUK in protein trafficking along the manchette and tail axoneme formation.