Aire regulates the transfer of antigen from mTECs to dendritic cells for induction of thymic tolerance.

To investigate the role of Aire in thymic selection, we examined the cellular requirements for generation of ovalbumin (OVA)-specific CD4 and CD8 T cells in mice expressing OVA under the control of the rat insulin promoter. Aire deficiency reduced the number of mature single-positive OVA-specific CD4(+) or CD8(+) T cells in the thymus, independent of OVA expression. Importantly, it also contributed in 2 ways to OVA-dependent negative selection depending on the T-cell type. Aire-dependent negative selection of OVA-specific CD8 T cells correlated with Aire-regulated expression of OVA. By contrast, for OVA-specific CD4 T cells, Aire affected tolerance induction by a mechanism that operated independent of the level of OVA expression, controlling access of antigen presenting cells to medullary thymic epithelial cell (mTEC)-expressed OVA. This study supports the view that one mechanism by which Aire controls thymic negative selection is by regulating the indirect presentation of mTEC-derived antigens by thymic dendritic cells. It also indicates that mTECs can mediate tolerance by direct presentation of Aire-regulated antigens to both CD4 and CD8 T cells.

Aire-deficient C57BL/6 mice mimicking the common human 13-base pair deletion mutation present with only a mild autoimmune phenotype.

Autoimmune regulator (AIRE) is an important transcription regulator that mediates a role in central tolerance via promoting the "promiscuous" expression of tissue-specific Ags in the thymus. Although several mouse models of Aire deficiency have been described, none has analyzed the phenotype induced by a mutation that emulates the common 13-bp deletion in human APECED (autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy) by disrupting the first plant homeodomain in exon 8. Aire-deficient mice with a corresponding mutation showed some disturbance of the medullary epithelial compartment, but at the phenotypic level their T cell compartment appeared relatively normal in the thymus and periphery. An increase in the number of activated T cells was evident, and autoantibodies against several organs were detected. At the histological level, lymphocytic infiltration of several organs indicated the development of autoimmunity, although symptoms were mild and the quality of life for Aire-deficient mice appeared equivalent to wild-type littermates, with the exception of male infertility. Vbeta and CDR3 length analysis suggested that each Aire-deficient mouse developed its own polyclonal autoimmune repertoire. Finally, given the prevalence of candidiasis in APECED patients, we examined the control of infection with Candida albicans in Aire-deficient mice. No increase in disease susceptibility was found for either oral or systemic infection. These observations support the view that additional genetic and/or environmental factors contribute substantially to the overt nature of autoimmunity associated with Aire mutations, even for mutations identical to those found in humans with APECED.

An integrated genetic and functional analysis of the role of type II transmembrane serine proteases (TMPRSSs) in hearing loss.

Building on our discovery that mutations in the transmembrane serine protease, TMPRSS3, cause nonsyndromic deafness, we have investigated the contribution of other TMPRSS family members to the auditory function. To identify which of the 16 known TMPRSS genes had a strong likelihood of involvement in hearing function, three types of biological evidence were examined: 1) expression in inner ear tissues; 2) location in a genomic interval that contains a yet unidentified gene for deafness; and 3) evaluation of hearing status of any available Tmprss knockout mouse strains. This analysis demonstrated that, besides TMPRSS3, another TMPRSS gene was essential for hearing and, indeed, mice deficient for Hepsin (Hpn) also known as Tmprss1 exhibited profound hearing loss. In addition, TMPRSS2, TMPRSS5, and CORIN, also named TMPRSS10, showed strong likelihood of involvement based on their inner ear expression and mapping position within deafness loci PKSR7, DFNB24, and DFNB25, respectively. These four TMPRSS genes were then screened for mutations in affected members of the DFNB24 and DFNB25 deafness families, and in a cohort of 362 sporadic deaf cases. This large mutation screen revealed numerous novel sequence variations including three potential pathogenic mutations in the TMPRSS5 gene. The mutant forms of TMPRSS5 showed reduced or absent proteolytic activity. Subsequently, TMPRSS genes with evidence of involvement in deafness were further characterized, and their sites of expression were determined. Tmprss1, 3, and 5 proteins were detected in spiral ganglion neurons. Tmprss3 was also present in the organ of Corti. TMPRSS1 and 3 proteins appeared stably anchored to the endoplasmic reticulum membranes, whereas TMPRSS5 was also detected at the plasma membrane. Collectively, these results provide evidence that TMPRSS1 and TMPRSS3 play and TMPRSS5 may play important and specific roles in hearing.

Mice deficient for the type II transmembrane serine protease, TMPRSS1/hepsin, exhibit profound hearing loss.

Defective proteolysis has been implicated in hearing loss through the discovery of mutations causing autosomal recessive nonsyndromic deafness in a type II transmembrane serine protease gene, TMPRSS3. To investigate their physiological function and the contribution of this family of proteases to the auditory function, we analyzed the hearing status of mice deficient for hepsin, also known as TMPRSS1. These mice exhibited profound hearing loss with elevated hearing thresholds compared with their heterozygous and wild-type littermates. Their cochleae showed abnormal tectorial membrane development, reduction in fiber compaction in the peripheral portion of the auditory nerve, and decreased expression of the myelin proteins myelin basic protein and myelin protein zero. In addition, reduced level of the large conductance voltage- and Ca(2+)-activated K(+) channel was detected in the sensory hair cells of Tmprss1-null mice. We examined thyroid hormone levels in Tmprss1-deficient mice, as similar cochlear defects have been reported in animal models of hypothyroidism, and found significantly reduced free thyroxine levels. These data show that TMPRSS1 is required for normal auditory function. Hearing impairment present in Tmprss1-null mice is characterized by a combination of various structural, cellular, and molecular abnormalities that are likely to affect different cochlear processes.