Alternative splicing regulates adaptor protein binding, trafficking, and activity of the Vps10p domain receptor SorCS2 in neuronal development.

The Vps10p domain receptor SorCS2 is crucial for the development and function of the nervous system and essential for brain-derived neurotrophic factor (BDNF)-induced changes in neuronal morphology and plasticity. SorCS2 regulates the subcellular trafficking of the BDNF signaling receptor TrkB as well as selected neurotransmitter receptors in a manner that is dependent on the SorCS2 intracellular domain (ICD). However, the cellular machinery and adaptor protein (AP) interactions that regulate receptor trafficking via the SorCS2 ICD are unknown. We here identify four splice variants of human SorCS2 differing in the insertion of an acidic cluster motif and/or a serine residue within the ICD. We show that each variant undergoes posttranslational proteolytic processing into a one- or two-chain receptor, giving rise to eight protein isoforms, the expression of which differs between neuronal and nonneuronal tissues and is affected by cellular stressors. We found that the only variants without the serine were able to rescue BDNF-induced branching of SorCS2 knockout hippocampal neurons, while variants without the acidic cluster showed increased interactions with clathrin-associated APs AP-1, AP-2, and AP-3. Using yeast two-hybrid screens, we further discovered that all variants bound dynein light chain Tctex-type 3; however, only variants with an acidic cluster motif bound kinesin light chain 1. Accordingly, splice variants showed markedly different trafficking properties and localized to different subcellular compartments. Taken together, our findings demonstrate the existence of eight functional SorCS2 isoforms with differential capacity for interactions with cytosolic ligands dynein light chain Tctex-type 3 and kinesin light chain 1, which potentially allows cell-type specific SorCS2 trafficking and BDNF signaling.

Loss of SORCS2 is Associated with Neuronal DNA Double-Strand Breaks.

SORCS2 is one of five proteins that constitute the Vps10p-domain receptor family. Members of this family play important roles in cellular processes linked to neuronal survival, differentiation and function. Genetic and functional studies implicate SORCS2 in cognitive function, as well as in neurodegenerative and psychiatric disorders. DNA damage and DNA repair deficits are linked to ageing and neurodegeneration, and transient neuronal DNA double-strand breaks (DSBs) also occur as a result of neuronal activity. Here, we report a novel role for SORCS2 in DSB formation. We show that SorCS2 loss is associated with elevated DSB levels in the mouse dentate gyrus and that knocking out SORCS2 in a human neuronal cell line increased Topoisomerase IIß-dependent DSB formation and reduced neuronal viability. Neuronal stimulation had no impact on levels of DNA breaks in vitro, suggesting that the observed differences may not be the result of aberrant neuronal activity in these cells. Our findings are consistent with studies linking the VPS10 receptors and DNA damage to neurodegenerative conditions.

Loss of BDNF or its receptors in three mouse models has unpredictable consequences for anxiety and fear acquisition.

BDNF-induced signaling is essential for the development of the central nervous system and critical for plasticity in adults. Mature BDNF signals through TrkB, while its precursor proBDNF employs p75(NTR), resulting in activation of signaling cascades with opposite effects on neuronal survival, growth cone decisions, and synaptic plasticity. Accordingly, variations in the genes encoding BDNF and its receptors sometimes have opposing influences in psychiatric disorders, and despite the vast literature, consensus is lacking about the behavioral consequences of disrupting the activity of the BDNF system in mice. To dissect the behavioral traits affected by dysfunctional BDNF/TrkB vs. proBDNF/p75(NTR) activity, we studied Bdnf(+/-), Ntrk2(+/-), and Ngfr(-/-) mice in parallel with respect to exploratory behavior, anxiety, startle, and fear acquisition. Our data reveal that the effect of proBDNF/BDNF and its receptors on behavior is more complex than expected. Strikingly, receptor-deficient mice displayed increased risk-taking behavior in the open field and elevated plus maze, whereas lack of proBDNF/BDNF had the opposite effect on mouse behavior. On the other hand, although TrkB signaling is instrumental for acquisition of fear memory in an inhibitory avoidance experiment, lack of p75(NTR) or proBDNF/BDNF conferred increased memory in this task. Importantly, none of the genotypes displayed any deficits in startle reflex, indicating unimpaired response to shock. The combined data illustrate an apparent paradox in the role of the BDNF system in controlling complex behavior and suggest that the individual components may also engage independently in separate signaling pathways.

PCSK9 deficiency alters brain lipid composition without affecting brain development and function.

PCSK9 induces lysosomal degradation of the low-density lipoprotein (LDL) receptor (LDLR) in the liver, hereby preventing removal of LDL cholesterol from the circulation. Accordingly, PCSK9 inhibitory antibodies and siRNA potently reduce LDL cholesterol to unprecedented low levels and are approved for treatment of hypercholesterolemia. In addition, PCSK9 inactivation alters the levels of several other circulating lipid classes and species. Brain function is critically influenced by cholesterol and lipid composition. However, it remains unclear how the brain is affected long-term by the reduction in circulating lipids as achieved with potent lipid lowering therapeutics such as PCSK9 inhibitors. Furthermore, it is unknown if locally expressed PCSK9 affects neuronal circuits through regulation of receptor levels. We have studied the effect of lifelong low peripheral cholesterol levels on brain lipid composition and behavior in adult PCSK9 KO mice. In addition, we studied the effect of PCSK9 on neurons in culture and in vivo in the developing cerebral cortex. We found that PCSK9 reduced LDLR and neurite complexity in cultured neurons, but neither PCSK9 KO nor overexpression affected cortical development in vivo. Interestingly, PCSK9 deficiency resulted in changes of several lipid classes in the adult cortex and cerebellum. Despite the observed changes, PCSK9 KO mice had unchanged behavior compared to WT controls. In conclusion, our findings demonstrate that altered PCSK9 levels do not compromise brain development or function in mice, and are in line with clinical trials showing that PCSK9 inhibitors have no adverse effects on cognitive function.

Altered dopaminergic firing pattern and novelty response underlie ADHD-like behavior of SorCS2-deficient mice.

Attention deficit hyperactivity disorder (ADHD) is the most frequently diagnosed neurodevelopmental disorder worldwide. Affected individuals present with hyperactivity, inattention, and cognitive deficits and display a characteristic paradoxical response to drugs affecting the dopaminergic system. However, the underlying pathophysiology of ADHD and how this relates to dopaminergic transmission remains to be fully understood. Sorcs2-/- mice uniquely recapitulate symptoms reminiscent of ADHD in humans. Here, we show that lack of SorCS2 in mice results in lower sucrose intake, indicating general reward deficits. Using in-vivo recordings, we further find that dopaminergic transmission in the ventral tegmental area (VTA) is shifted towards a more regular firing pattern with marked reductions in the relative occurrence of irregular firing in Sorcs2-/- mice. This was paralleled by abnormal acute behavioral responses to dopamine receptor agonists, suggesting fundamental differences in dopaminergic circuits and indicating a perturbation in the balance between the activities of the postsynaptic dopamine receptor DRD1 and the presynaptic inhibitory autoreceptor DRD2. Interestingly, the hyperactivity and drug response of Sorcs2-/- mice were markedly affected by novelty. Taken together, our findings show how loss of a candidate ADHD-risk gene has marked effects on dopaminergic circuit function and the behavioral response to the environment.

Reduced Alcohol Seeking and Withdrawal Symptoms in Mice Lacking the BDNF Receptor SorCS2.

Alcohol use disorder (AUD) is characterized by repetitive and uncontrolled intake of alcohol with severe consequences for affected individuals, their families and society as a whole. Numerous studies have implicated brain-derived neurotrophic factor (BDNF) activity in the neurobiology underlying AUD. The BDNF signaling mechanism is complex and depends on two receptor systems, TrkB and p75NTR, which appear to have opposite effects on alcohol seeking behavior in animal models. We recently discovered that the sortilin-related receptor SorCS2 forms complexes with both TrkB and p75NTR and is important for BDNF activity in the developing and adult CNS. Moreover, the SORCS2 gene was recently identified as the top association signal for severity of alcohol withdrawal symptoms. Hence, we speculated that SorCS2 deficient mice would have an altered response to alcohol. The role of SorCS2 in the acute and adapted response to alcohol was therefore investigated by comparing SorCS2 knockout (Sorcs2-/- ) mice to wild type (WT) mice in three paradigms modeling alcohol sensitivity and consumption; alcohol-induced conditioned place preference, two-bottle choice test as well as the behavioral response to alcohol withdrawal. We found that, when compared to the WT mice, (I) Sorcs2-/- mice displayed complete lack of alcohol-induced place preference, (II) when given free choice between water and alcohol, Sorcs2-/- mice consumed less alcohol, and (III) Sorcs2-/- mice showed no handling-induced convulsion in response to alcohol withdrawal following extended alcohol exposure. Taken together, these results show that lack of the alcohol withdrawal risk gene Sorcs2 results in abnormal behavioral response to alcohol in mice. Consequently, SorCS2 may play an important role in the molecular pathways underlying AUD and complications associated with alcohol withdrawal.

Exploring the sortilin related receptor, SorLA, in depression.

BACKGROUND: Studies of individual biomarkers for depression have shown insufficient sensitivity and specificity for clinical use, and most likely combinations of biomarkers may provide a better signature. The sorting-related receptor with A-type repeats (SorLA) is a well-studied pathogenic factor for Alzheimer's. SorLA belongs to the Vps10p domain receptor family, which also encompasses sortilin and SorCS1-3. All family members have been implicated in neurological and mental disorders. Notably, the SORCS3 gene is genome-wide significantly associated with depression and serum protein levels of sortilin are reduced in depressed individuals. SorLA regulates the activity of neurotrophic factors and cytokines and we hence speculated that SorLA might be implicated in depression. METHODS: Serum SorLA levels were measured in two well-defined clinical samples using ELISA. Generalized linear models were used in the statistical analyses. RESULTS: We identified a multivariate model to discriminate depressed individuals from healthy controls. Interestingly, the model consisted of serum SorLA levels and additional four predictors: previous depressive episode, stressful life events, serum levels of sortilin and VEGF. However, as an isolated factor, we observed no significant difference in SorLA levels between 140 depressed individuals and 140 healthy controls. Nevertheless, we observed a significant increase in SorLA levels following 12 weeks of treatment with nortriptyline, but not escitalopram. LIMITATIONS: The number of biomarkers included in the multivariate model for depression and lack of replication limit our study. CONCLUSIONS: Our results suggest SorLA as one of five factors that in combination may support the depression diagnosis, but not as an individual biomarker for depression or treatment response.