Age-Related Changes in Thymic Central Tolerance.

Thymic epithelial cells (TECs) and hematopoietic antigen presenting cells (HAPCs) in the thymus microenvironment provide essential signals to self-reactive thymocytes that induce either negative selection or generation of regulatory T cells (Treg), both of which are required to establish and maintain central tolerance throughout life. HAPCs and TECs are comprised of multiple subsets that play distinct and overlapping roles in central tolerance. Changes that occur in the composition and function of TEC and HAPC subsets across the lifespan have potential consequences for central tolerance. In keeping with this possibility, there are age-associated changes in the cellular composition and function of T cells and Treg. This review summarizes changes in T cell and Treg function during the perinatal to adult transition and in the course of normal aging, and relates these changes to age-associated alterations in thymic HAPC and TEC subsets.

Polycomb Repressive Complex 2 is essential for development and maintenance of a functional TEC compartment.

Thymic epithelial cells (TEC) are essential for thymocyte differentiation and repertoire selection. Despite their indispensable role in generating functional T cells, the molecular mechanisms that orchestrate TEC development from endodermal progenitors in the third pharyngeal pouch (3rd PP) are not fully understood. We recently reported that the T-box transcription factor TBX1 negatively regulates TEC development. Although initially expressed throughout the 3rd PP, Tbx1 becomes downregulated in thymus-fated progenitors and when ectopically expressed impairs TEC progenitor proliferation and differentiation. Here we show that ectopic Tbx1 expression in thymus fated endoderm increases expression of Polycomb repressive complex 2 (PRC2) target genes in TEC. PRC2 is an epigenetic modifier that represses gene expression by catalyzing trimethylation of lysine 27 on histone H3. The increased expression of PRC2 target genes suggests that ectopic Tbx1 interferes with PRC2 activity and implicates PRC2 as an important regulator of TEC development. To test this hypothesis, we used Foxn1Cre to delete Eed, a PRC2 component required for complex stability and function in thymus fated 3rd PP endoderm. Proliferation and differentiation of fetal and newborn TEC were disrupted in the conditional knockout (EedCKO) mutants leading to severely dysplastic adult thymi. Consistent with PRC2-mediated transcriptional silencing, the majority of differentially expressed genes (DEG) were upregulated in EedCKO TEC. Moreover, a high frequency of EedCKO DEG overlapped with DEG in TEC that ectopically expressed Tbx1. These findings demonstrate that PRC2 plays a critical role in TEC development and suggest that Tbx1 expression must be downregulated in thymus fated 3rd PP endoderm to ensure optimal PRC2 function.

Tissue specific expression of SG2NA is regulated by differential splicing, RNA editing and differential polyadenylation.

SG2NA belongs to a three member Striatin subfamily of WD-40 repeat superfamily. It has multiple protein-protein interaction domains that are involved in the assembly of supra-molecular signaling complexes. Earlier we had demonstrated that there are at least five variants of SG2NA, generated by alternative splicing. We now demonstrate that a 52kDa novel variant is generated by the editing of the transcript for the 82kDa isoform. The 52kDa protein is abundant in mouse tissues but it is barely present in immortalized cells, suggesting its role in cell differentiation. Besides splicing and editing, expression of SG2NAs in tissues is also regulated by differential polyadenylation and mRNA/protein stability. Further, the longer UTR is seen only in the brain mRNA from 1month old mouse and 8-10day old chick embryo. Like alternative splicing, differential polyadenylation of Sg2na transcripts is also conserved in evolution. Taken together, these results suggest a highly versatile and dynamic mode of regulation of SG2NA with potential implications in tissue development.

Among the three striatin family members, SG2NA was first to arise during evolution.

Striatin, SG2NA, and zinedin constitute a three-member subfamily of WD-40 repeat proteins. They are found only in metazoans and are likely to have scaffolding functions. Apart from WD-40 repeats, they also have a caveolin-binding motif, a coiled-coil structure, and a calmodulin-binding domain. This paper focuses on the analysis of their evolution as a paradigm of understanding the metazoan scaffolds. Each member of the family forms distinct phylogenetic clusters, wherein striatins, SG2NAs, and zinedins have 13, 10, and 9 conserved motifs, respectively. Furthermore, two of those motifs each in striatin and in zinedin and three in SG2NA are exclusive for the respective subfamily. Of those exclusive motifs for SG2NA, two encompass the caveolin-binding and coiled-coiled domains. Collectively, they show the presence of 11 conserved motifs, suggestive of convergence of individual motifs and creation of patterns. A prokaryotic WD-40 repeat motif pCM-I was found only in the corresponding domain of SG2NA but not in other family members. It is thus hypothesized that striatin family members have evolved from bacteria, and SG2NA was the first member to arise.

WD-40 repeat protein SG2NA has multiple splice variants with tissue restricted and growth responsive properties.

SG2NA is a member of the striatin family of WD-40 repeat proteins with potential scaffolding functions. It was originally identified as a tumor antigen with increased expression during S to G2 phase of cell cycle. We report here that mouse SG2NA has at least five novel splice variants of which two are devoid of the carboxyl terminal WD-40 repeats. The variants of SG2NA are generated by alternative splicing at the exon 7-9 regions and differ in their expression profiles in various tissues tested. While the 83, 78, 38 and 35 kDa variants are present in both brain and heart, the 87 kDa form is brain specific. Also, the expression of 35 kDa variant is more in neonatal than in adult tissues. Western analysis suggests that the SG2NA isoforms differentially respond to growth stimuli. Upon serum stimulation, while the 35 kDa variant is increased, the 78 kDa form is diminished. Splicing variation of SG2NA is conserved in metazoan evolution. In embryonic chicken there are at least four variants of which one is present in brain but absent in heart. Taken together, splicing variation of SG2NA might have some critical roles in differentiation and maturation in metazoan cells.