Subclavian artery aneurysmal rupture and left internal mammary artery extravasation secondary to advanced Marfan syndrome.

This case highlights the unusual life-threatening findings found in a patient with Marfan syndrome (MFS) in the emergency department setting. MFS is a rare autosomal dominant disease that affects 1 in 3000-5000 individuals and has a highly variable range of clinical severity. This case is a 63-year-old male with COPD, scoliosis, aortic and mitral valve replacements on warfarin, and MFS who presented with acute onset hemoptysis, tachypnea, and oxygen saturation of 77% on 4 l nasal cannula. Emergent chest computed tomography angiography (CTA) revealed both a contained rupture of a left subclavian artery aneurysm and active extravasation from his left internal mammary artery (LIMA) into his left chest. The patient was on warfarin and reversed with IV vitamin K and prothrombin complex concentrate. Vascular surgery emergently took the patient to the operating room for embolization of his LIMA and stenting of the contained ruptured left subclavian artery aneurysm. The patient was discharged home one month after admission. This case report illustrates the potential severe sequelae of MFS and the importance of rapid recognition by emergency physicians. An expanded understanding of the pathophysiology of MFS has resulted in great advancement in medical therapies and lifestyle modification and thus has significantly prolonged life expectancy in these patients. Increased awareness and familiarity will facilitate continued high-quality management and treatment by emergency physicians.

Chloroplast SRP43 acts as a chaperone for glutamyl-tRNA reductase, the rate-limiting enzyme in tetrapyrrole biosynthesis.

Assembly of light-harvesting complexes requires synchronization of chlorophyll (Chl) biosynthesis with biogenesis of light-harvesting Chl a/b-binding proteins (LHCPs). The chloroplast signal recognition particle (cpSRP) pathway is responsible for transport of nucleus-encoded LHCPs in the stroma of the plastid and their integration into the thylakoid membranes. Correct folding and assembly of LHCPs require the incorporation of Chls, whose biosynthesis must therefore be precisely coordinated with membrane insertion of LHCPs. How the spatiotemporal coordination between the cpSRP machinery and Chl biosynthesis is achieved is poorly understood. In this work, we demonstrate a direct interaction between cpSRP43, the chaperone that mediates LHCP targeting and insertion, and glutamyl-tRNA reductase (GluTR), a rate-limiting enzyme in tetrapyrrole biosynthesis. Concurrent deficiency for cpSRP43 and the GluTR-binding protein (GBP) additively reduces GluTR levels, indicating that cpSRP43 and GBP act nonredundantly to stabilize GluTR. The substrate-binding domain of cpSRP43 binds to the N-terminal region of GluTR, which harbors aggregation-prone motifs, and the chaperone activity of cpSRP43 efficiently prevents aggregation of these regions. Our work thus reveals a function of cpSRP43 in Chl biosynthesis and suggests a striking mechanism for posttranslational coordination of LHCP insertion with Chl biosynthesis.

Two distinct sites of client protein interaction with the chaperone cpSRP43.

Integral membrane proteins are prone to aggregation and misfolding in aqueous environments and therefore require binding by molecular chaperones during their biogenesis. Chloroplast signal recognition particle 43 (cpSRP43) is an ATP-independent chaperone required for the biogenesis of the most abundant class of membrane proteins, the light-harvesting chlorophyll a/b-binding proteins (LHCPs). Previous work has shown that cpSRP43 specifically recognizes an L18 loop sequence conserved among LHCP paralogs. However, how cpSRP43 protects the transmembrane domains (TMDs) of LHCP from aggregation was unclear. In this work, alkylation-protection and site-specific cross-linking experiments found that cpSRP43 makes extensive contacts with all the TMDs in LHCP. Site-directed mutagenesis identified a class of cpSRP43 mutants that bind tightly to the L18 sequence but are defective in chaperoning full-length LHCP. These mutations mapped to hydrophobic surfaces on or near the bridging helix and the ß-hairpins lining the ankyrin repeat motifs of cpSRP43, suggesting that these regions are potential sites for interaction with the client TMDs. Our results suggest a working model for client protein interactions in this membrane protein chaperone.

Permeability transition pore-dependent and PARP-mediated depletion of neuronal pyridine nucleotides during anoxia and glucose deprivation.

Exposure of rat cortical neurons to combined oxygen and glucose deprivation results in loss of NAD(P)H autofluorescence that is only partially reversible following restoration of oxygen and glucose, suggesting catabolism of pyridine nucleotides. This study tested the hypothesis that metabolic inhibition caused by cyanide-induced chemical anoxia plus glucose deprivation promotes both release of mitochondrial NAD(H) in response to opening of the permeability transition pore (PTP) and NAD(P)(H) degradation through activation of poly (ADP-ribose) polymerase (PARP). The NAD(P)H autofluorescence of rat neonatal cortical neurons was monitored during and following acute (10-30 min) exposure to the respiratory inhibitor, cyanide, in the absence and presence of glucose. Because nitric oxide-derived peroxynitrite is a known activator of PARP, we additionally assessed the effect of a nitric oxide generating agent on the NAD(P)H autofluorescence response to chemical anoxia plus glucose deprivation. Cyanide induced a rapid increase in autofluorescence, followed by a steady decline promoted by the presence of nitric oxide. This decline was primarily due to NAD(H) catabolism, as verified by measurements of total NAD(H) present in cellular extracts. Catabolism was partially blocked by an inhibitor of PARP, by a PTP inhibitor, and by either glucose or pyruvate as a source of reducing power. Overall, data suggest that metabolic, oxidative, and nitrosative stress during in vitro neuronal anoxia and glucose deprivation result in release of mitochondrial pyridine nucleotides in response to PTP opening and rapid, extensive NAD(H) degradation mediated by PARP activation. These events may contribute to the metabolic dysfunction that occurs in vivo during cerebral ischemia and reperfusion and therefore represent prime targets for neuroprotection.

Dodecamer assembly of a metazoan AAA+ chaperone couples substrate extraction to refolding.

Ring-forming AAA+ chaperones solubilize protein aggregates and protect organisms from proteostatic stress. In metazoans, the AAA+ chaperone Skd3 in the mitochondrial intermembrane space (IMS) is critical for human health and efficiently refolds aggregated proteins, but its underlying mechanism is poorly understood. Here, we show that Skd3 harbors both disaggregase and protein refolding activities enabled by distinct assembly states. High-resolution structures of Skd3 hexamers in distinct conformations capture ratchet-like motions that mediate substrate extraction. Unlike previously described disaggregases, Skd3 hexamers further assemble into dodecameric cages in which solubilized substrate proteins can attain near-native states. Skd3 mutants defective in dodecamer assembly retain disaggregase activity but are impaired in client refolding, linking the disaggregase and refolding activities to the hexameric and dodecameric states of Skd3, respectively. We suggest that Skd3 is a combined disaggregase and foldase, and this property is particularly suited to meet the complex proteostatic demands in the mitochondrial IMS.

Chloroplast SRP43 autonomously protects chlorophyll biosynthesis proteins against heat shock.

The assembly of light-harvesting chlorophyll-binding proteins (LHCPs) is coordinated with chlorophyll biosynthesis during chloroplast development. The ATP-independent chaperone known as chloroplast signal recognition particle 43 (cpSRP43) mediates post-translational LHCP targeting to the thylakoid membrane and also participates in tetrapyrrole biosynthesis (TBS). How these distinct actions of cpSRP43 are controlled has remained unclear. Here, we demonstrate that cpSRP43 effectively protects several TBS proteins from heat-induced aggregation and enhances their stability during leaf greening and heat shock. While the substrate-binding domain of cpSRP43 is sufficient for chaperoning LHCPs, the stabilization of TBS clients requires the chromodomain 2 of the protein. Strikingly, cpSRP54-which activates cpSRP43's LHCP-targeted function-inhibits the chaperone activity of cpSRP43 towards TBS proteins. High temperature weakens the interaction of cpSRP54 with cpSRP43, thus freeing cpSRP43 to interact with and protect the integrity of TBS proteins. Our data indicate that the temperature sensitivity of the cpSRP43-cpSRP54 complex enables cpSRP43 to serve as an autonomous chaperone for the thermoprotection of TBS proteins.

A Disorder-to-Order Transition Activates an ATP-Independent Membrane Protein Chaperone.

The 43 kDa subunit of the chloroplast signal recognition particle, cpSRP43, is an ATP-independent chaperone essential for the biogenesis of the light harvesting chlorophyll-binding proteins (LHCP), the most abundant membrane protein family on earth. cpSRP43 is activated by a stromal factor, cpSRP54, to more effectively capture and solubilize LHCPs. The molecular mechanism underlying this chaperone activation is unclear. Here, a combination of hydrogen-deuterium exchange, electron paramagnetic resonance, and NMR spectroscopy experiments reveal that a disorder-to-order transition of the ankyrin repeat motifs in the substrate binding domain of cpSRP43 drives its activation. An analogous coil-to-helix transition in the bridging helix, which connects the ankyrin repeat motifs to the cpSRP54 binding site in the second chromodomain, mediates long-range allosteric communication of cpSRP43 with its activating binding partner. Our results provide a molecular model to explain how the conformational dynamics of cpSRP43 enables regulation of its chaperone activity and suggest a general mechanism by which ATP-independent chaperones with cooperatively folding domains can be regulated.

Testing geographical pathways of speciation in a recent island radiation.

Determining the mode, or geographical context, of speciation is a critical first step to understanding the evolutionary mechanisms that cause new species to arise. In this study, we estimated phylogenetic relationships in the cerasina species group of the Hawaiian cricket genus Laupala (Orthoptera: Gryllidae) to test competing phylogeographical hypotheses and thus infer the mode of speciation. A previous phylogenetic result based on nuclear sequence data suggested that populations of L. cerasina on the Big Island of Hawaii are the result of two independent colonizations from Maui, implying parallel speciation and convergent song evolution, and contradicting systematic hypotheses based on behavioural and morphological data. We used amplified fragment length polymorphisms to investigate further the relationships among species and populations in the cerasina species group. Results of these analyses provide a robust estimate of phylogenetic relationships and support the phylogeographical history indicated by behavioural and morphological data.