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Diabetes mellitus (DM) is a chronic metabolic disorder characterized by inappropriate hyperglycemia, which causes endothelial dysfunction and peripheral neuropathy, ultimately leading to multiple complications. One prevalent complication is diabetic erectile dysfunction (ED), which is more severe and more resistant to treatment than nondiabetic ED. The serum glycoprotein leucine-rich ɑ-2-glycoprotein 1 (LRG1) is a modulator of TGF-β-mediated angiogenesis and has been proposed as a biomarker for a variety of diseases, including DM. Here, we found that the adhesion GPCR latrophilin-2 (LPHN2) is a TGF-β-independent receptor of LRG1. By interacting with LPHN2, LRG1 promotes both angiogenic and neurotrophic processes in mouse tissue explants under hyperglycemic conditions. Preclinical studies in a diabetic ED mouse model showed that LRG1 administration into the penile tissue, which exhibits significantly increased LPHN2 expression, fully restores erectile function by rescuing vascular and neurological abnormalities. Further investigations revealed that PI3K, AKT, and NF-κB p65 constitute the key intracellular signaling pathway of the LRG1/LPHN2 axis, providing important mechanistic insights into LRG1-mediated angiogenesis and nerve regeneration in DM. Our findings suggest that LRG1 can be a potential new therapeutic option for treating aberrant peripheral blood vessels and neuropathy associated with diabetic complications, such as diabetic ED.

The serum glycoprotein leucine-rich ɑ-2-glycoprotein 1 (LRG1), primarily produced by hepatocytes and neutrophils, is a multifunctional protein that can modulate various signalling cascades, mainly TGFβ signalling. Serum LRG1 and neutrophil-derived LRG1 have different molecular weights due to differences in glycosylation, but what impact the differential glycan composition in LRG1 has on its cellular function is largely unknown. We previously reported that LRG1 can promote both angiogenic and neurotrophic processes under hyperglycemic conditions by interacting with LPHN2. Here, we determined the crystal structure of LRG1, identifying the horseshoe-like solenoid structure of LRG1 and its four N-glycosylation sites. In addition, our biochemical and cell-biological analysis found that de-glycosylation of LRG1, particularly the removal of glycans on N325, is critical for high-affinity binding of LRG1 to LPHN2, thereby promoting LRG1/LPHN2-mediated angiogenic and neurotrophic processes in mouse tissue explants, even under normal glucose conditions. Moreover, intracavernous administration of de-glycosylated LRG1 in a diabetic mouse model ameliorated vascular and neurological abnormalities and restored erectile function. Collectively, these data indicate a novel role of LRG1’s glycans as molecular switches that can tune the range of LRG1’s cellular functions, particularly the LRG1/LPHN2 signalling axis.

The retinal pigment epithelium (RPE) forms a monolayer sheet separating the retina and choroid in vertebrate eyes. The polarized nature of RPE is maintained by distributing membrane proteins differentially along apico-basal axis. We found the distributions of these proteins differ in embryonic, post-natal, and mature mouse RPE, suggesting developmental regulation of protein trafficking. Thus, we deleted tumor susceptibility gene 101 (Tsg101), a key component of endosomal sorting complexes required for transport (ESCRT), in embryonic and mature RPE to determine whether ESCRT-mediated endocytic protein trafficking correlated with the establishment and maintenance of RPE polarity. Loss of Tsg101 severely disturbed the polarity of RPE, which forms irregular aggregates exhibiting non-polarized distribution of cell adhesion proteins and activation of epidermal growth factor receptor signaling. These findings suggest that ESCRT-mediated protein trafficking is essential for the development and maintenance of RPE cell polarity.

International audience; OTX2 (orthodenticle homeobox 2) haplodeficiency causes diverse defects in mammalian visual systems ranging from retinal dysfunction to anophthalmia. We find that the retinal dystrophy of Otx2+/GFP heterozygous knockin mice is mainly due to the loss of bipolar cells and consequent deficits in retinal activity. Among bipolar cell types, OFF-cone bipolar subsets, which lack autonomous Otx2 gene expression but receive Otx2 proteins from photoreceptors, degenerate most rapidly in Otx2+/GFP mouse retinas, suggesting a neuroprotective effect of the imported Otx2 protein. In support of this hypothesis, retinal dystrophy in Otx2+/GFP mice is prevented by intraocular injection of Otx2 protein, which localizes to the mitochondria of bipolar cells and facilitates ATP synthesis as a part of mitochondrial ATP synthase complex. Taken together, our findings demonstrate a mitochondrial function for Otx2 and suggest a potential therapeutic application of OTX2 protein delivery in human retinal dystrophy.

Retinal ganglion cell (RGC) axons of binocular animals cross the midline at the optic chiasm (OC) to grow toward their synaptic targets in the contralateral brain. Ventral anterior homeobox 1 (Vax1) plays an essential role in the development of the OC by regulating RGC axon growth in a non-cell autonomous manner. In this study, we identify an unexpected function of Vax1 that is secreted from ventral hypothalamic cells and diffuses to RGC axons, where it promotes axonal growth independent of its transcription factor activity. We demonstrate that Vax1 binds to extracellular sugar groups of the heparan sulfate proteoglycans (HSPGs) located in RGC axons. Both Vax1 binding to HSPGs and subsequent penetration into the axoplasm, where Vax1 activates local protein synthesis, are required for RGC axonal growth. Together, our findings demonstrate that Vax1 possesses a novel RGC axon growth factor activity that is critical for the development of the mammalian binocular visual system. DOI: http://dx.doi.org/10.7554/eLife.02671.001, eLife digest We see the world around us when light bounces off of objects and hits the retina at the back of our eyes. This triggers electrical signals in neurons called retinal ganglion cells (RGCs), which have long structures called axons that extend out from the retina and into the parts of the brain where the signals are interpreted. As the axons grow, various ‘guidance’ molecules direct the axons to the correct part of the brain. One molecule that is important for the growth of retinal ganglion cells' axons is a protein called Vax1. This protein is a transcription factor and binds to DNA to control how and when the molecular templates used to make proteins are made—a process called transcription. Vax1 is not produced in retinal ganglion cells, but it does control the extension of these cells' axons into part of the brain called the ventral hypothalamus. In this study, the axons cross to the other side of the brain by forming a structure called optic chiasm. Humans and mice lacking Vax1 are unable to develop the optic chiasm, and the axons of their retinal ganglion cells do not reach their targets in the brain. These defects were thought to occur because the guidance molecules whose transcription is normally controlled by Vax1 were not produced in the correct amounts when Vax1 is absent. Kim et al. now challenge this view by creating a mutant version of Vax1 that cannot bind to DNA or regulate the transcription of other proteins. Retinal ganglion cell axons could still grow correctly when they were put close to cells expressing this version of the Vax1 protein. This contradicts a hypothesis that Vax1 supports axonal growth by transcribing guidance molecules. Kim et al. followed up these results by examining developing mice and reached the unexpected conclusion that Vax1 is secreted from cells in the ventral hypothalamus and binds to a type of sugar molecule found on the surface of the axons. Once bound, Vax1 can enter the axons where it appears to stimulate the production of proteins inside axons, which helps the axons to grow. These findings reveal unconventional functions for Vax1 that occur in addition to its role as a transcription factor. Vax1 is known to regulate the development of several structures in the brain, so the work of Kim et al. also raises new questions about how Vax1 controls these processes. DOI: http://dx.doi.org/10.7554/eLife.02671.002