Diabetes, obesity, and cancer affect upward of 15% of the world's population. Interestingly, all three diseases juxtapose dysregulated intracellular signaling with altered metabolic state. Exactly which genetic factors define stable metabolic set points in vivo remains poorly understood. Here, we show that hedgehog signaling rewires cellular metabolism. We identify a cilium-dependent Smo-Ca(2+)-Ampk axis that triggers rapid Warburg-like metabolic reprogramming within minutes of activation and is required for proper metabolic selectivity and flexibility. We show that Smo modulators can uncouple the Smo-Ampk axis from canonical signaling and identify cyclopamine as one of a new class of "selective partial agonists," capable of concomitant inhibition of canonical and activation of noncanonical hedgehog signaling. Intriguingly, activation of the Smo-Ampk axis in vivo drives robust insulin-independent glucose uptake in muscle and brown adipose tissue. These data identify multiple noncanonical endpoints that are pivotal for rational design of hedgehog modulators and provide a new therapeutic avenue for obesity and diabetes.

Mature focal adhesions and fibrillar adhesions act as anchorage sites for vimentin filaments, with plectin isoform 1f being the crucial linker protein. Plectin serves as a nucleation and assembly center for the de novo formation of vimentin networks. Anchored vimentin creates a resilient cage-like core structure that affects cell shape.

Plectin is a large, 500-kDa, intermediate filament (IF)-associated protein. It acts as a cytoskeletal crosslinker and signaling scaffold, affecting mechanical as well as dynamic properties of the cytoskeleton. As a member of the plakin family of cytolinker proteins, plectin has a multidomain structure that is responsible for its vast binding portfolio. It not only binds to all types of IFs, actin filaments and microtubules, but also to transmembrane receptors, proteins of the subplasma membrane protein skeleton, components of the nuclear envelope, and several kinases with known roles in migration, proliferation, and energy metabolism of cells. Due to alternative splicing, plectin is expressed as various isoforms with differing N-terminal heads that dictate their differential subcellular targeting. Through specific interactions with other proteins at their target sites and their ability to bind to all types of IFs, plectin molecules provide strategically located IF anchorage sites within the cytoplasm of cells. In this review, we will present an overview of the structural features and functional properties of plectin and discuss recent progress in defining the role of its isoforms in stress-prone tissues and the implicated diseases, with focus on skin, skeletal muscle, and Schwann cells of peripheral nerve.

Plectin, a giant multifunctional cytolinker protein, plays a crucial role in stabilizing and orchestrating intermediate filament networks in cells. Mutations in the human plectin gene result in multiple diseases manifesting with muscular dystrophy, skin blistering, and signs of neuropathy. The most common disease caused by plectin deficiency is epidermolysis bullosa simplex (EBS)-MD, a rare autosomal-recessive skin blistering disorder with late-onset muscular dystrophy. EBS-MD patients and plectin-deficient mice display pathologic desmin-positive protein aggregates, degenerated myofibrils, and mitochondrial abnormalities, the hallmarks of myofibrillar myopathies. In addition to EBS-MD, plectin mutations have been shown to cause EBS-MD with a myasthenic syndrome, limb-girdle muscular dystrophy type 2Q, EBS with pyloric atresia, and EBS-Ogna. This review focuses on clinical and pathological manifestations of these plectinopathies. It addresses especially plectin's role in skeletal muscle, where a loss of muscle fiber integrity and profound changes of myofiber cytoarchitecture are observed in its absence. Furthermore, the highly complex genetic and molecular structure of plectin is discussed; a high number of differentially spliced exons give rise to a variety of different isoforms, which fulfill distinct functions in different cell types and tissues. Plectin's abilities to act as a dynamic organizer of intermediate filament networks and to interact with a multitude of different interaction partners are the basis for its function as a scaffolding platform for proteins involved in signaling. Finally, the article addresses a series of genetically manipulated mouse lines that were generated to serve as powerful models to study functional and molecular consequences of plectin gene defects.

Mutations of the human desmin gene on chromosome 2q35 cause autosomal dominant, autosomal recessive and sporadic forms of protein aggregation myopathies and cardiomyopathies. We generated R349P desmin knock-in mice, which harbor the ortholog of the most frequently occurring human desmin missense mutation R350P. These mice develop age-dependent desmin-positive protein aggregation pathology, skeletal muscle weakness, dilated cardiomyopathy, as well as cardiac arrhythmias and conduction defects. For the first time, we report the expression level and subcellular distribution of mutant versus wild-type desmin in our mouse model as well as in skeletal muscle specimens derived from human R350P desminopathies. Furthermore, we demonstrate that the missense-mutant desmin inflicts changes of the subcellular localization and turnover of desmin itself and of direct desmin-binding partners. Our findings unveil a novel principle of pathogenesis, in which not the presence of protein aggregates, but disruption of the extrasarcomeric intermediate filament network leads to increased mechanical vulnerability of muscle fibers. These structural defects elicited at the myofiber level finally impact the entire organ and subsequently cause myopathy and cardiomyopathy.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-014-1363-2) contains supplementary material, which is available to authorized users.

Abbreviations used in this paper: ATPS, ATP synthase; cyt c , cytochrome c ; IF, intermediate fi lament; mito-PAGFP, mitochondrial matrix -targeted photoactivatable GFP; NAO, 10-N -nonyl-acridine orange; P1b, plectin 1b; RACK1, receptor for activated C kinase 1; ROI, region of interest; vim, vimentin; wt, wild type.The online version of this paper contains supplemental material. IntroductionMitochondria perform a multitude of cellular activities that are essential for a cell ' s life and death. There is evidence that mitochondrial morphology and distribution depend on interactions with the cytoskeleton, although the molecular mechanisms involved are hardly understood ( Toivola et al., 2005 ;Anesti and Scorrano, 2006 ). A connection of mitochondria with intermediate fi laments (IFs) was suggested some 25 years ago ( Toh et al., 1980 ), and several IF proteins have been associated with mitochondrial functions since then. Mutations in the neurofi lament protein NF-L gene have been shown to affect mitochondrial distribution ( Perez-Olle et al., 2005 ), and an aberrant mitochondrial distribution in keratinocytes was observed in some patients with epidermolysis bullosa simplex caused by mutations in keratins 5 and 14 genes ( Uttam et al., 1996 ). Furthermore, the ablation of desmin in the mouse results in characteristic alterations in distribution, number, morphology, and respiratory activity of mitochondria (for review see Capetanaki et al., 2007 ). The question of whether the interaction between IFs and mitochondria occurs directly or is mediated by linker proteins remains to be solved.The highly versatile IF-based cytolinker protein plectin ( Wiche, 1998 ) would be an interesting candidate for mediating the interactions between IFs and mitochondria. The versatility of plectin is largely caused by complex splicing events in the N-terminal region of its gene that give rise to 11 alternatively spliced isoforms containing different fi rst exons (1 -1j;Elliott et al., 1997 ;Fuchs et al., 1999 ). The expression patterns of these isoforms are cell type -dependent, and some of the expressed variants have been shown to differ in their subcellular localization ( Rezniczek et al., 2003 ). By forced expression in fi broblasts, isoform plectin 1b (P1b) was found to be specifi cally targeted to mitochondria ( Rezniczek et al., 2003 ). Here, we analyzed the mode of P1b -mitochondrion interaction and show that this interaction affects the shape and network formation of mitochondria. Results and discussionMitochondrion-associated P1b is an outer membrane -anchored protein facing the cytosol First, we analyzed the mode of P1b interaction with mitochondria and the topology of its molecular subdomains. After subcellular fractionation of mouse fi broblasts, the distribution of P1b was found to be very similar to that of genuine mitochondrial proteins but different from other plectin isoforms (Fig. S1, A and B, available P lectin is a versatile intermediate fi lament (IF) -bound cytolinker protein with a variety of differentially splice...

Organometallic metal(arene) anticancer agents require ligand exchange for their anticancer activity and this is generally believed to confer low selectivity for potential cellular targets. However, using an integrated proteomics-based target-response profiling approach as a potent hypothesis-generating procedure, we found an unexpected target selectivity of a ruthenium(arene) pyridinecarbothioamide (plecstatin) for plectin, a scaffold protein and cytolinker, which was validated in a plectin knock-out model in vitro. Plectin targeting shows potential as a strategy to inhibit tumor invasiveness as shown in cultured tumor spheroids while oral administration of plecstatin-1 to mice reduces tumor growth more efficiently in the invasive B16 melanoma than in the CT26 colon tumor model.

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