Background: Cyclophilins harbor ill-defined chaperone and prolyl isomerase activities toward physiological substrates. Results: Nonoverlapping chaperone or prolyl isomerase activity loss of Ran-binding protein 2 (Ranbp2) cyclophilin domain triggers unique impairments of proteostasis in distinct cell types and ubiquitin-proteasome system. Conclusion: Ranbp2 cyclophilin subdomains present discriminating physiological activities toward substrates or regulation of ubiquitin-proteasome system. Significance: Ranbp2-mediated mechanistic links in proteostasis with physiological and therapeutic relevance are uncovered.The immunophilins, cyclophilins, catalyze peptidyl cis-trans prolyl-isomerization (PPIase), a rate-limiting step in protein folding and a conformational switch in protein function. Cyclophilins are also chaperones. Noncatalytic mutations affecting the only cyclophilins with known but distinct physiological substrates, the Drosophila NinaA and its mammalian homolog, cyclophilin-B, impair opsin biogenesis and cause osteogenesis imperfecta, respectively. However, the physiological roles and substrates of most cyclophilins remain unknown. It is also unclear if PPIase and chaperone activities reflect distinct cyclophilin properties. To elucidate the physiological idiosyncrasy stemming from potential cyclophilin functions, we generated mice lacking endogenous Ran-binding protein-2 (Ranbp2) and expressing bacterial artificial chromosomes of Ranbp2 with impaired C-terminal chaperone and with (Tg-Ranbp2 WT-HA ) or without PPIase activities (Tg-Ranbp2 R2944A-HA ). The transgenic lines exhibit unique effects in proteostasis. Either line presents selective deficits in M-opsin biogenesis with its accumulation and aggregation in cone photoreceptors but without proteostatic impairment of two novel Ranbp2 cyclophilin partners, the cytokine-responsive effectors, STAT3/STAT5. Peptidyl cis-trans-prolyl isomerization is a rate-limiting step in protein folding (1-3). The catalysis of the cis-trans interconversion of the peptidyl-prolyl isomers is catalyzed by peptidylprolyl cis-trans isomerases (PPIase) 5 (4 -6). PPIases compose three families of structurally unrelated proteins, the cyclophilins (CyP), FK506-binding proteins (FKBP), and parvulins (7). * This work was supported, in whole or in part, by National Institutes of Health Grants EY019492, GM083165, and GM083165-03S1 (to P. A. F.), 2P30-EY005722 (to Duke University Eye Center), and 5P30NS061789 (to Duke Neurotransgenic Laboratory). This work was also supported by the Foundation Fighting CyPs and FKBPs are designated also as immunophilins, because they mediate immunosuppression (8,9). This effect is achieved by a gain-of-function mechanism upon binding of the immunosuppressive metabolites, cyclosporin A (CsA) or FK506, to the PPIase active site and formation of a ternary complex with the serine/threonine phosphatase, calcineurin, whose sequestration and inhibition prevents the dephosphorylation and activation of the nuclear factor for activation of T-cells (9 -12). Howe...

The pathogenic drivers of sporadic and familial motor neuron disease (MND), such amyotrophic lateral sclerosis (ALS), are unknown. MND impairs the Ran GTPase cycle, which controls nucleocytoplasmic transport, ribostasis and proteostasis; however, cause-effect mechanisms of Ran GTPase modulators in motoneuron pathobiology have remained elusive. The cytosolic and peripheral nucleoporin Ranbp2 is a crucial regulator of the Ran GTPase cycle and of the proteostasis of neurological disease-prone substrates, but the roles of Ranbp2 in motoneuron biology and disease remain unknown. This study shows that conditional ablation of Ranbp2 in mouse Thy1 motoneurons causes ALS syndromes with hypoactivity followed by hindlimb paralysis, respiratory distress and, ultimately, death. These phenotypes are accompanied by: a decline in the nerve conduction velocity, free fatty acids and phophatidylcholine of the sciatic nerve; a reduction in the g-ratios of sciatic and phrenic nerves; and hypertrophy of motoneurons. Furthermore, Ranbp2 loss disrupts the nucleocytoplasmic partitioning of the import and export nuclear receptors importin β and exportin 1, respectively, Ran GTPase and histone deacetylase 4. Whole-transcriptome, proteomic and cellular analyses uncovered that the chemokine receptor Cxcr4, its antagonizing ligands Cxcl12 and Cxcl14, and effector, latent and activated Stat3 all undergo early autocrine and proteostatic deregulation, and intracellular sequestration and aggregation as a result of Ranbp2 loss in motoneurons. These effects were accompanied by paracrine and autocrine neuroglial deregulation of hnRNPH3 proteostasis in sciatic nerve and motoneurons, respectively, and post-transcriptional downregulation of metalloproteinase 28 in the sciatic nerve. Mechanistically, our results demonstrate that Ranbp2 controls nucleocytoplasmic, chemokine and metalloproteinase 28 signaling, and proteostasis of substrates that are crucial to motoneuronal homeostasis and whose impairments by loss of Ranbp2 drive ALS-like syndromes.

Background: Ranbp2 and its Ran-GTP-binding domains' roles in RPE survival/function, a multidisease target, are elusive. Results: RPE undergoes degeneration, disruptions of proteostasis of Ranbp2 partners, and blood-retinal barrier upon Ranbp2 ablation. Impairment of selective Ran-GTP-binding domains of Ranbp2 suffices to promote RPE degeneration. Conclusion: Ran GTPase regulation by Ranbp2 is vital to RPE. Significance: Ranbp2-dependent targets/mechanisms with therapeutic potential in RPE degeneration are uncovered. The retinal pigment epithelium (RPE) 6 is a critical component of the blood-retinal barrier and maintains retinal homeostasis by filtering damaging light, nourishing the neural retina, replenishing the 11-cis-retinal chromophore to photoreceptors, and phagocytizing damaged (photo-oxidized) outer segments of photoreceptors (1-3). RPE dysfunction underlies disparate diseases promoting RPE degeneration, such as atrophic and neovascular age-related macular degeneration (4 -6), * This work was supported, in whole or in part, by National Institutes of Health Grants EY019492, GM083165, and GM083165-04S1 (to P. A. F.), 2P30-EY005722 (to the Duke University Eye Center), and 5P30NS061789 (to the Duke Neurotransgenic Laboratory). This work was also supported by the Foundation Fighting Blindness bestrophinopathies (7), and various forms of retinal dystrophies (e.g. retinitis pigmentosa, Leber congenital amaurosis) (8 -12). The cause-effect mechanisms of RPE degeneration are not understood, but they appear to arise from the interplay of multifactorial events impinging on heterogeneous genetic predispositions, noxious insults, and senescence that culminate with cytotoxicity to the RPE (13-21). Retinal pigment epithelium (RPE) degeneration underpins diseases triggered by disparate genetic lesions, noxious insultsEmerging studies indicate that the modular and pleiotropic Ranbp2 (Ran (Ras-related nuclear protein)-binding protein 2) plays critical and cell type-dependent roles in cell survival, proliferation, or functions upon intrinsic or extrinsic pathological stressors. For example, haploinsufficiency of Ranbp2 in an inbred genetic background confers neuroprotection to photoreceptor degeneration upon photo-oxidative stress and modulates glucose tolerance (22-24), whereas it increases the susceptibility of mice to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-elicited Parkinsonism (25) or carcinogenesis (26). Interestingly, RANBP2 is also a substrate for degradation by PARKIN (27,28), whose impairment causes familial and sporadic Parkinson (29 -31) or multisite oncogenesis (32,33). Further, Ranbp2 loss in cones photoreceptors also elicits non-autonomous death of healthy rod photoreceptors (34), whereas asymptomatic mutations in human RANBP2 predispose the basal ganglia to acute necrotic damage (e.g. encephalopathy) upon exposure to various infectious agents (35-37). A parsimonious model of the multifold activities of multimodular Ranbp2 arises from structure-function analyses of Ranbp2, whereby the dynamic recruitme...