The 22q11.2 region regulates presynaptic gene-products linked to schizophrenia.

It is unclear how the 22q11.2 deletion predisposes to psychiatric disease. To study this, we generated induced pluripotent stem cells from deletion carriers and controls and utilized CRISPR/Cas9 to introduce the heterozygous deletion into a control cell line. Here, we show that upon differentiation into neural progenitor cells, the deletion acted in trans to alter the abundance of transcripts associated with risk for neurodevelopmental disorders including autism. In excitatory neurons, altered transcripts encoded presynaptic factors and were associated with genetic risk for schizophrenia, including common and rare variants. To understand how the deletion contributed to these changes, we defined the minimal protein-protein interaction network that best explains gene expression alterations. We found that many genes in 22q11.2 interact in presynaptic, proteasome, and JUN/FOS transcriptional pathways. Our findings suggest that the 22q11.2 deletion impacts genes that may converge with psychiatric risk loci to influence disease manifestation in each deletion carrier.

Activity-dependent alpha-cleavage of nectin-1 is mediated by a disintegrin and metalloprotease 10 (ADAM10).

Nectin-1 is known to undergo ectodomain shedding by alpha-secretase and subsequent proteolytic processing by gamma-secretase. How secretase-mediated cleavage of nectin-1 is regulated in neuronal cells and how nectin-1 cleavage affects synaptic adhesion is poorly understood. We have investigated alpha-and gamma-secretase-mediated processing of nectin-1 in primary cortical neurons and identified which protease acts as a alpha-secretase. We report here that NMDA receptor activation, but not stimulation of AMPA or metabotropic glutamate receptors, resulted in robust alpha- and gamma-secretase cleavage of nectin-1 in mature cortical neurons. Cleavage of nectin-1 required influx of Ca(2+) through the NMDA receptor, and activation of calmodulin, but was not dependent on calcium/calmodulin-dependent protein kinase II (CaMKII) activation. We found that ADAM10 is the major secretase responsible for nectin-1 ectodomain cleavage in neurons and the brain. These observations suggest that alpha- and gamma-secretase processing of nectin-1 is a Ca(2+)/calmodulin-regulated event that occurs under conditions of activity-dependent synaptic plasticity and ADAM10 and gamma-secretase are responsible for these cleavage events.

Brain-derived neurotrophic factor signaling in the HVC is required for testosterone-induced song of female canaries.

Testosterone-induced singing in songbirds is thought to involve testosterone-dependent morphological changes that include angiogenesis and neuronal recruitment into the HVC, a central part of the song control circuit. Previous work showed that testosterone induces the production of vascular endothelial growth factor (VEGF) and its receptor (VEGFR2 tyrosine kinase), which in turn leads to an upregulation of brain-derived neurotrophic factor (BDNF) production in HVC endothelial cells. Here we report for the first time that systemic inhibition of the VEGFR2 tyrosine kinase is sufficient to block testosterone-induced song in adult female canaries, despite sustained androgen exposure and the persistence of the effects of testosterone on HVC morphology. Expression of exogenous BDNF in HVC, induced locally by in situ transfection, reversed the VEGFR2 inhibition-mediated blockade of song development, thereby restoring the behavioral phenotype associated with androgen-induced song. The VEGFR2-inhibited, BDNF-treated females developed elaborate male-like song that included large syllable repertoires and high syllable repetition rates, features known to attract females. Importantly, although functionally competent new neurons were recruited to HVC after testosterone treatment, the time course of neuronal addition appeared to follow BDNF-induced song development. These findings indicate that testosterone-associated VEGFR2 activity is required for androgen-induced song in adult songbirds and that the behavioral effects of VEGFR2 inhibition can be rescued by BDNF within the adult HVC.

Testosterone-induced matrix metalloproteinase activation is a checkpoint for neuronal addition to the adult songbird brain.

Testosterone-induced neuronal addition to the adult songbird vocal control center, HVC, requires the androgenic induction of vascular endothelial growth factor (VEGF), followed by VEGF-stimulated angiogenesis. The expanded vasculature acts as a source of BDNF, which supports the immigration of new neurons from the overlying ventricular zone. In tumorigenesis, a similar process of adult angiogenesis is regulated by matrix metalloproteinase (MMP) activity, in particular that of the gelatinases. We therefore investigated the role of the gelatinases in neuronal addition to the HVC of adult female canaries. In situ zymography of the caudal forebrain revealed that testosterone-induced perivascular gelatinase activity that was most prominent in HVC. High-resolution gels revealed distinct MMP activities that comigrated with MMP2 and MMP9, and PCR cloning yielded MMP2 and MMP9 orthologues of 1465 and 1044 bp, respectively. Quantitative PCR revealed that HVC MMP2 mRNA levels doubled within 8 d of testosterone, whereas MMP9 transcript levels were stable. Moreover, isolated adult canary forebrain endothelial cells secreted MMP2, and VEGF substantially increased endothelial MMP2 gelatinase activity. To assess the importance of androgen-regulated, VEGF-induced MMP2 to adult angiogenesis and neurogenesis, we treated testosterone-implanted females with the gelatinase inhibitor SB-3CT. In situ zymography confirmed that SB-3CT suppressed gelatinase activity in HVC, and histological analysis revealed that SB-3CT-treated birds exhibited a decreased endothelial mitotic index and substantially diminished neuronal recruitment to HVC. These data suggest that the androgenic induction of endothelial MMP2 is a critical regulator of neuronal addition to the adult HVC, and as such comprises an important regulatory step in adult neurogenesis.

Altered Abeta formation and long-term potentiation in a calsenilin knock-out.

Calsenilin has been identified as a presenilin-binding protein, a transcription factor regulating dynorphin expression, and a beta-subunit of Kv4 channels and could, thus, be a multifunctional protein. To study these functions of calsenilin in vivo and to determine the neuroanatomical expression pattern of calsenilin, we generated mice with a disruption of the calsenilin gene by the targeted insertion of the beta-galactosidase gene. We found that calsenilin expression (as represented by beta-galactosidase activity) is very restricted but overlaps better with that of presenilins and Kv4 channels than with dynorphin, suggesting that calsenilin may regulate presenilin and Kv4 channels in brain. Abeta peptide levels are reduced in calsenilin knock-out mice, demonstrating that calsenilin affects presenilin-dependent gamma-cleavage in vivo. Furthermore, long-term potentiation (LTP) in dentate gyrus of hippocampus, in which calsenilin is strongly and selectively expressed, is enhanced in calsenilin knock-out mice. This enhancement of LTP coincides with a downregulation of the Kv4 channel-dependent A-type current and can be mimicked in wild-type animals by a Kv4 channel blocker. The data presented here show that lack of calsenilin affects both Abeta formation and the A-type current. We suggest that these effects are separate events, caused by a common mechanism possibly involving protein transport.