Polymers and inflammation: disease mechanisms of the serpinopathies

Members of the serpin (serine proteinase inhibitor) superfamily play a central role in the control of inflammatory, coagulation, and fibrinolytic cascades. Point mutations that cause abnormal conformational transitions in these proteins can trigger disease. Recent work has defined three pathways by which these conformers cause tissue damage. Here, we describe how these three mechanisms can be integrated into a new model of the pathogenesis of emphysema caused by mutations in the serpin α1-antitrypsin

Why has it been so difficult to prove the efficacy of alpha-1-antitrypsin replacement therapy? Insights from the study of disease pathogenesis

Alpha-1-antitrypsin is the most abundant circulating protease inhibitor. It is mainly produced by the liver and secreted into the circulation where it acts to prevent excessive proteolytic damage in the lungs by the enzyme neutrophil elastase. The most common severe deficiency allele is the Z mutation, which causes the protein to self-associate into ordered polymers. These polymers accumulate within hepatocytes to cause liver damage. The resulting lack of circulating α1-antitrypsin predisposes the Z homozygote to proteolytic lung damage and emphysema. Other pathways may also contribute to the development of lung disease. In particular, polymers of Z α1-antitrypsin can form within the lung where they act as a pro-inflammatory stimulus that may exacerbate protease-mediated lung damage. Researchers recognized in the 1980s that plasma α1-antitrypsin levels could be restored by intravenous infusions of purified human protein. Alpha-1-antitrypsin replacement therapy was introduced in 1987 but subsequent clinical trials have produced conflicting results, and to date there remains no widely accepted clinical evidence of the efficacy of α1-antitrypsin replacement therapy. This review addresses our current understanding of disease pathogenesis in α1-antitrypsin deficiency and questions why this treatment in isolation may not be effective. In particular it discusses the possible role of α1-antitrypsin polymers in exacerbating intrapulmonary inflammation and attenuating the efficacy of α1-antitrypsin replacement therapy

Genetic susceptibility.

Why only 20% of smokers develop clinically relevant chronic obstructive pulmonary disease (COPD) was a puzzle for many years. Now, epidemiologic studies point clearly toward a large heritable component. The combination of genome-wide association studies and candidate gene analysis is helping to identify those genetic variants responsible for an individual's susceptibility to developing COPD. In this review, the current data implicating specific loci and genes in the pathogenesis of COPD are examined

Endoplasmic reticulum polymers impair luminal protein mobility and sensitize to cellular stress in alpha1-antitrypsin deficiency.

UNLABELLED: Point mutants of alpha1 -antitrypsin (α1AT) form ordered polymers that are retained as inclusions within the endoplasmic reticulum (ER) of hepatocytes in association with neonatal hepatitis, cirrhosis, and hepatocellular carcinoma. These inclusions cause cell damage and predispose to ER stress in the absence of the classical unfolded protein response (UPR). The pathophysiology underlying this ER stress was explored by generating cell models that conditionally express wild-type (WT) α1AT, two mutants that cause polymer-mediated inclusions and liver disease (E342K [the Z allele] and H334D) and a truncated mutant (Null Hong Kong; NHK) that induces classical ER stress and is removed by ER-associated degradation. Expression of the polymeric mutants resulted in gross changes in the ER luminal environment that recapitulated the changes observed in liver sections from individuals with PI*ZZ α1AT deficiency. In contrast, expression of NHK α1AT caused electron lucent dilatation and expansion of the ER throughout the cell. Photobleaching microscopy in live cells demonstrated a decrease in the mobility of soluble luminal proteins in cells that express E342K and H334D α1AT, when compared to those that express WT and NHK α1AT (0.34 ± 0.05, 0.22 ± 0.03, 2.83 ± 0.30, and 2.84 ± 0.55 μm(2) /s, respectively). There was no effect on protein mobility within ER membranes, indicating that cisternal connectivity was not disrupted. Polymer expression alone was insufficient to induce the UPR, but the resulting protein overload rendered cells hypersensitive to ER stress induced by either tunicamycin or glucose depletion. CONCLUSION: Changes in protein diffusion provide an explanation for the cellular consequences of ER protein overload in mutants that cause inclusion body formation and α1AT deficiency

Therapeutic target-site variability in α1-antitrypsin characterized at high resolution

The intrinsic propensity of [alpha]1-antitrypsin to undergo conformational transitions from its metastable native state to hyperstable forms provides a motive force for its antiprotease function. However, aberrant conformational change can also occur via an intermolecular linkage that results in polymerization. This has both loss-of-function and gain-of-function effects that lead to deficiency of the protein in human circulation, emphysema and hepatic cirrhosis. One of the most promising therapeutic strategies being developed to treat this disease targets small molecules to an allosteric site in the [alpha]1-antitrypsin molecule. Partial filling of this site impedes polymerization without abolishing function. Drug development can be improved by optimizing data on the structure and dynamics of this site. A new 1.8 Å resolution structure of [alpha]1-antitrypsin demonstrates structural variability within this site, with associated fluctuations in its upper and lower entrance grooves and ligand-binding characteristics around the innermost stable enclosed hydrophobic recess. These data will allow a broader selection of chemotypes and derivatives to be tested in silico and in vitro when screening and developing compounds to modulate conformational change to block the pathological mechanism while preserving function

Endoplasmic reticulum dysfunction in neurological disease.

Endoplasmic reticulum (ER) dysfunction might have an important part to play in a range of neurological disorders, including cerebral ischaemia, sleep apnoea, Alzheimer's disease, multiple sclerosis, amyotrophic lateral sclerosis, the prion diseases, and familial encephalopathy with neuroserpin inclusion bodies. Protein misfolding in the ER initiates the well studied unfolded protein response in energy-starved neurons during stroke, which is relevant to the toxic effects of reperfusion. The toxic peptide amyloid β induces ER stress in Alzheimer's disease, which leads to activation of similar pathways, whereas the accumulation of polymeric neuroserpin in the neuronal ER triggers a poorly understood ER-overload response. In other neurological disorders, such as Parkinson's and Huntington's diseases, ER dysfunction is well recognised but the mechanisms by which it contributes to pathogenesis remain unclear. By targeting components of these signalling responses, amelioration of their toxic effects and so the treatment of a range of neurodegenerative disorders might become possible

The intracellular accumulation of polymeric neuroserpin explains the severity of the dementia FENIB

Familial encephalopathy with neuroserpin inclusion bodies (FENIB) is an autosomal dominant dementia that is characterized by the retention of polymers of neuroserpin as inclusions within the endoplasmic reticulum (ER) of neurons. We have developed monoclonal antibodies that detect polymerized neuroserpin and have used COS-7 cells, stably transfected PC12 cell lines and transgenic Drosophila melanogaster to characterize the cellular handling of all four mutant forms of neuroserpin that cause FENIB. We show a direct correlation between the severity of the disease-causing mutation and the accumulation of neuroserpin polymers in cell and fly models of the disease. Moreover, mutant neuroserpin causes locomotor deficits in the fly allowing us to demonstrate a direct link between polymer accumulation and neuronal toxicity

The TRiC/CCT chaperone is implicated in Alzheimer's disease based on patient GWAS and an RNAi screen in Aβ-expressing Caenorhabditis elegans.

The human Aβ peptide causes progressive paralysis when expressed in the muscles of the nematode worm, C. elegans. We have exploited this model of Aβ toxicity by carrying out an RNAi screen to identify genes whose reduced expression modifies the severity of this locomotor phenotype. Our initial finding was that none of the human orthologues of these worm genes is identical with the genome-wide significant GWAS genes reported to date (the "white zone"); moreover there was no identity between worm screen hits and the longer list of GWAS genes which included those with borderline levels of significance (the "grey zone"). This indicates that Aβ toxicity should not be considered as equivalent to sporadic AD. To increase the sensitivity of our analysis, we then considered the physical interactors (+1 interactome) of the products of the genes in both the worm and the white+grey zone lists. When we consider these worm and GWAS gene lists we find that 4 of the 60 worm genes have a +1 interactome overlap that is larger than expected by chance. Two of these genes form a chaperonin complex, the third is closely associated with this complex and the fourth gene codes for actin, the major substrate of the same chaperonin

Decoupling of net community and export production on submesoscales in the Sargasso Sea

Author Posting. © American Geophysical Union, 2015. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Global Biogeochemical Cycles 29 (2015): 1266–1282, doi:10.1002/2014GB004913.Determinations of the net community production (NCP) in the upper ocean and the particle export production (EP) should balance over long time and large spatial scales. However, recent modeling studies suggest that a horizontal decoupling of flux-regulating processes on submesoscales (≤10 km) could lead to imbalances between individual determinations of NCP and EP. Here we sampled mixed-layer biogeochemical parameters and proxies for NCP and EP during 10, high-spatial resolution (~2 km) surface transects across strong physical gradients in the Sargasso Sea. We observed strong biogeochemical and carbon flux variability in nearly all transects. Spatial coherence among measured biogeochemical parameters within transects was common but rarely did the same parameters covary consistently across transects. Spatial variability was greater in parameters associated with higher trophic levels, such as chlorophyll in >5.0 µm particles, and variability in EP exceeded that of NCP in nearly all cases. Within sampling transects, coincident EP and NCP determinations were uncorrelated. However, when averaged over each transect (30 to 40 km in length), we found NCP and EP to be significantly and positively correlated (R = 0.72, p = 0.04). Transect-averaged EP determinations were slightly smaller than similar NCP values (Type-II regression slope of 0.93, standard deviation = 0.32) but not significantly different from a 1:1 relationship. The results show the importance of appropriate sampling scales when deriving carbon flux budgets from upper ocean observations.NASA Ocean Carbon and Biogeochemistry program Grant Number: NNX11AL94G; WHOI Postdoctoral Scholar fellowship; NASA ACE Grant Number: NNX12AJ25G; NSF Grant Number: OCE-07523662016-02-2