Seven Kinds of Intermediate Filament Networks in the Cytoplasm of Polarized Cells: Structure and Function

Intermediate filaments (IFs) are involved in many important physiological functions, such as the distribution of organelles, signal transduction, cell polarity and gene regulation. However, little information exists on the structure of the IF networks performing these functions. We have clarified the existence of seven kinds of IF networks in the cytoplasm of diverse polarized cells: an apex network just under the terminal web, a peripheral network lying just beneath the cell membrane, a granule-associated network surrounding a mass of secretory granules, a Golgi-associated network surrounding the Golgi apparatus, a radial network locating from the perinuclear region to the specific area of the cell membrane, a juxtanuclear network surrounding the nucleus, and an entire cytoplasmic network. In this review, we describe these seven kinds of IF networks and discuss their biological roles

Linking cytoarchitecture to metabolism: sarcolemma-associated plectin affects glucose uptake by destabilizing microtubule networks in mdx myofibers

BACKGROUND: Duchenne muscular dystrophy (DMD) is one of the most frequent forms of muscular disorders. It is caused by the absence of dystrophin, a core component of the sarcolemma-associated junctional complex that links the cytoskeleton to the extracellular matrix. We showed previously that plectin 1f (P1f), one of the major muscle-expressed isoforms of the cytoskeletal linker protein plectin, accumulates at the sarcolemma of DMD patients as well as of mdx mice, a widely studied animal model for DMD.Based on plectin's dual role as structural protein and scaffolding platform for signaling molecules, we speculated that the dystrophic phenotype observed after loss of dystrophin was caused, at least to some extent, by excess plectin. Thus, we hypothesized that elimination of plectin expression in mdx skeletal muscle, while probably resulting in an overall more severe phenotype, may lead to a partial phenotype rescue. In particular, we wanted to assess whether excess sarcolemmal plectin contributes to the dysregulation of sugar metabolism in mdx myofibers. METHODS: We generated plectin/dystrophin double deficient (dKO) mice by breeding mdx with conditional striated muscle-restricted plectin knockout (cKO) mice. The phenotype of these mice was comparatively analyzed with that of mdx, cKO, and wild-type mice, focusing on structural integrity and dysregulation of glucose metabolism. RESULTS: We show that the accumulation of plectin at the sarcolemma of mdx muscle fibers hardly compensated for their loss of structural integrity. Instead, it led to an additional metabolic deficit by impairing glucose uptake. While dKO mice suffered from an overall more severe form of muscular dystrophy compared to mdx or plectin-deficient mice, sarcolemmal integrity as well as glucose uptake of their myofibers were restored to normal levels upon ablation of plectin. Furthermore, microtubule (MT) networks in intact dKO myofibers, including subsarcolemmal areas, were found to be more robust than those in mdx mice. Finally, myotubes differentiated from P1f-overexpressing myoblasts showed an impairment of glucose transporter 4 translocation and a destabilization of MT networks. CONCLUSIONS: Based on these results we propose that sarcolemma-associated plectin acts as an antagonist of MT network formation in myofibers, thereby hindering vesicle-mediated (MT-dependent) transport of glucose transporter 4. This novel role of plectin throws a bridge between extra-sarcomeric cytoarchitecture and metabolism of muscle fibers. Our study thus provides new insights into pathomechanisms of plectinopathies and muscular dystrophies in general

Life-long course and molecular characterization of the original Dutch family with epidermolysis bullosa simplex with muscular dystrophy due to a homozygous novel plectin point mutation

Plectin is one of the largest and most versatile cytolinker proteins known. Cloned and sequenced in 1991, it was later shown to have nonsense mutations in recessive epidermolysis bullosa with muscular dystrophy. A dominant mutation in the gene was found to cause epidermolysis bullosa simplex Ogna without muscular dystrophy. Here we report the DNA sequencing of the plectin gene (PLEC1) in a Dutch family originally described in 1972 as having epidermolysis bullosa with muscular dystrophy. The results revealed homozygosity for a new plectin nonsense mutation at position 13187 and its specific 8q24 marker haplotype profile. Western blotting of cultured fibroblasts and immunofluorescence microscopy of skin biopsy confirm that the plectin protein expression is grossly reduced or absent. A summary of the life-long clinical course of the two affected brothers homozygous for the new E1914X mutation is given

Life-long course and molecular characterization of the original Dutch family with epidermolysis bullosa simplex with muscular dystrophy due to a homozygous novel plectin point mutation

Plectin is one of the largest and most versatile cytolinker proteins known. Cloned and sequenced in 1991, it was later shown to have nonsense mutations in recessive epidermolysis bullosa with muscular dystrophy. A dominant mutation in the gene was found to cause epidermolysis bullosa simplex Ogna without muscular dystrophy. Here we report the DNA sequencing of the plectin gene (PLEC1) in a Dutch family originally described in 1972 as having epidermolysis bullosa with muscular dystrophy. The results revealed homozygosity for a new plectin nonsense mutation at position 13187 and its specific 8q24 marker haplotype profile. Western blotting of cultured fibroblasts and immunofluorescence microscopy of skin biopsy confirm that the plectin protein expression is grossly reduced or absent. A summary of the life-long clinical course of the two affected brothers homozygous for the new E1914X mutation is given.</p

Plectin as a prognostic marker in non-metastatic oral squamous cell carcinoma

Background: Oral squamous cell carcinoma (OSCC) is associated with a poor 5-year survival rate. In general, patients diagnosed with small tumors have a fairly good prognosis, but some small tumors have an aggressive behavior leading to early death. There are at present no reliable prognostic biomarkers for oral cancers. Thus, to optimize treatment for the individual patient, there is a need for biomarkers that can predict tumor behavior. Method: In the present study the potential prognostic value of plectin was evaluated by a tissue microarray (TMA) based immunohistochemical analysis of primary tumor tissue obtained from a North Norwegian cohort of 115 patients diagnosed with OSCC. The expression of plectin was compared with clinicopathological variables and 5 year survival. Results: The statistical analysis revealed that low expression of plectin in the tumor cells predicted a favorable outcome for patients with non-metastatic disease (p = 0.008). Furthermore, the expression of plectin was found to correlate (p = 0.01) with the expression of uPAR, which we have previously found to be a potential prognostic marker for T1N0 tumors. Conclusions: Our results indicate that low expression of plectin predicts a favorable outcome for patients with non-metastatic OSCC and the expression level of plectin may therefore be used in the treatment stratification for patients with early stage disease

Keeping the Vimentin Network under Control: Cell–Matrix Adhesion–associated Plectin 1f Affects Cell Shape and Polarity of Fibroblasts

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

Computational Characterization of 3′ Splice Variants in the GFAP Isoform Family

Glial fibrillary acidic protein (GFAP) is an intermediate filament (IF) protein specific to central nervous system (CNS) astrocytes. It has been the subject of intense interest due to its association with neurodegenerative diseases, and because of growing evidence that IF proteins not only modulate cellular structure, but also cellular function. Moreover, GFAP has a family of splicing isoforms apparently more complex than that of other CNS IF proteins, consistent with it possessing a range of functional and structural roles. The gene consists of 9 exons, and to date all isoforms associated with 3′ end splicing have been identified from modifications within intron 7, resulting in the generation of exon 7a (GFAPδ/ε) and 7b (GFAPκ). To better understand the nature and functional significance of variation in this region, we used a Bayesian multiple change-point approach to identify conserved regions. This is the first successful application of this method to a single gene – it has previously only been used in whole-genome analyses. We identified several highly or moderately conserved regions throughout the intron 7/7a/7b regions, including untranslated regions and regulatory features, consistent with the biology of GFAP. Several putative unconfirmed features were also identified, including a possible new isoform. We then integrated multiple computational analyses on both the DNA and protein sequences from the mouse, rat and human, showing that the major isoform, GFAPα, has highly conserved structure and features across the three species, whereas the minor isoforms GFAPδ/ε and GFAPκ have low conservation of structure and features at the distal 3′ end, both relative to each other and relative to GFAPα. The overall picture suggests distinct and tightly regulated functions for the 3′ end isoforms, consistent with complex astrocyte biology. The results illustrate a computational approach for characterising splicing isoform families, using both DNA and protein sequences