The cell adhesion molecule BT-IgSF is essential for a functional blood-testis barrier and male fertility in mice.

The Ig-like cell adhesion molecule (IgCAM) BT-IgSF (brain- and testis-specific Ig superfamily protein) plays a major role in male fertility in mice. However, the molecular mechanism by which BT-IgSF supports fertility is unclear. Here, we found that it is localized in Sertoli cells at the blood-testis barrier (BTB) and at the apical ectoplasmic specialization. The absence of BT-IgSF in Sertoli cells in both global and conditional mouse mutants (i.e. AMHCre and Rosa26CreERT2 lines) resulted in male infertility, atrophic testes with vacuolation, azoospermia, and spermatogenesis arrest. Although transcripts of junctional proteins such as connexin43, ZO-1, occludin, and claudin11 were up-regulated in the absence of BT-IgSF, the functional integrity of the BTB was impaired, as revealed by injection of a BTB-impermeable component into the testes under in vivo conditions. Disruption of the BTB coincided with mislocalization of connexin43, which was present throughout the seminiferous epithelium and not restricted to the BTB as in wild-type tissues, suggesting impaired cell-cell communication in the BT-IgSF-KO mice. Because EM images revealed a normal BTB structure between Sertoli cells in the BT-IgSF-KO mice, we conclude that infertility in these mice is most likely caused by a functionally impaired BTB. In summary, our results indicate that BT-IgSF is expressed at the BTB and is required for male fertility by supporting the functional integrity of the BTB.

The IgCAM BT-IgSF (IgSF11) is essential for connexin43-mediated astrocyte-astrocyte coupling in mice.

The type I transmembrane protein BT-IgSF is predominantly localized in the brain and testes. It belongs to the CAR subgroup of Ig cell adhesion proteins, that are hypothesized to regulate connexin expression or localization. Here, we studied the putative link between BT-IgSF and connexins in astrocytes, ependymal cells and neurons of the mouse. Global knockout of BT-IgSF caused an increase in the clustering of connexin43 (Gja1), but not of connexin30 (Gjb6), on astrocytes and ependymal cells. Additionally, knockout animals displayed reduced expression levels of connexin43 protein in the cortex and hippocampus. Importantly, analysis of biocytin spread in hippocampal or cortical slices from mature mice of either sex revealed a decrease in astrocytic cell-cell coupling in the absence of BT-IgSF. Blocking either protein biosynthesis or proteolysis showed that the lysosomal pathway increased connexin43 degradation in astrocytes. Localization of connexin43 in subcellular compartments was not impaired in astrocytes of BT-IgSF mutants. In contrast to connexin43 the localization and expression of connexin36 (Gjd2) on neurons was not affected by the absence of BT-IgSF. Overall, our data indicate that the IgCAM BT-IgSF is essential for correct gap junction-mediated astrocyte-to-astrocyte cell communication.Significance Statement Astrocytes regulate a variety of physiological processes in the developing and adult brain that are essential for proper brain function. Astrocytes form extensive networks in the brain and communicate via gap junctions. Disruptions of gap junction coupling are found in several diseases such as neurodegeneration or epilepsy. Here, we demonstrate that the cell adhesion protein BT-IgSF is essential for gap junction mediated coupling between astrocytes in the cortex and hippocampus.

Lack of the Ig cell adhesion molecule BT-IgSF (IgSF11) induced behavioral changes in the open maze, water maze and resident intruder test.

The brain- and testis-specific Ig superfamily protein (BT-IgSF, also termed IgSF11) is a homotypic cell adhesion protein. In the nervous system, BT-IgSF regulates the stability of AMPA receptors in the membrane of cultured hippocampal neurons, modulates the connectivity of chandelier cells and controls gap junction-mediated astrocyte-astrocyte communication. Here, we performed behavioral tests in BT-IgSF-deficient mice. BT-IgSF-deficient mice were similar to control littermates with respect to their reflexes, motor coordination and gating, and associative learning. However, BT-IgSF-deficient mice displayed an increased tendency to stay in the central illuminated areas in the open field and O-Maze paradigms suggesting reduced anxiety or increased scotophobia (fear of darkness). Although BT-IgSF-deficient mice initially found the platform in the water maze their behavior was compromised when the platform was moved, indicating reduced behavioral flexibility. This deficit was overcome by longer training to improve their spatial memory. Furthermore, male BT-IgSF-deficient mice displayed increased aggression towards an intruder. Our results show that specific behaviors are modified by the lack of BT-IgSF and demonstrate a contribution of BT-IgSF to network functions.

The IgCAM CLMP regulates expression of Connexin43 and Connexin45 in intestinal and ureteral smooth muscle contraction in mice.

CAR-like membrane protein (CLMP), an immunoglobulin cell adhesion molecule (IgCAM), has been implicated in congenital short-bowel syndrome in humans, a condition with high mortality for which there is currently no cure. We therefore studied the function of CLMP in a Clmp-deficient mouse model. Although we found that the levels of mRNAs encoding Connexin43 or Connexin45 were not or were only marginally affected, respectively, by Clmp deficiency, the absence of CLMP caused a severe reduction of both proteins in smooth muscle cells of the intestine and of Connexin43 in the ureter. Analysis of calcium signaling revealed a disordered cell-cell communication between smooth muscle cells, which in turn induced an impaired and uncoordinated motility of the intestine and the ureter. Consequently, insufficient transport of chyme and urine caused a fatal delay to thrive, a high rate of mortality, and provoked a severe hydronephrosis in CLMP knockouts. Neurotransmission and the capability of smooth muscle cells to contract in ring preparations of the intestine were not altered. Physical obstructions were not detectable and an overall normal histology in the intestine as well as in the ureter was observed, except for a slight hypertrophy of smooth muscle layers. Deletion of Clmp did not lead to a reduced length of the intestine as shown for the human CLMP gene but resulted in gut malrotations. In sum, the absence of CLMP caused functional obstructions in the intestinal tract and ureter by impaired peristaltic contractions most likely due to a lack of gap-junctional communication between smooth muscle cells.