Cells, Volume 7, Issue 4 (April 2018) – 10 articles

The mammalian nuclear lamina proteins—prelamin A- and B-type lamins—are post-translationally modified by farnesylation, endoproteolysis, and carboxymethylation at a carboxy-terminal CAAX (C, cysteine; a, aliphatic amino acid; X, any amino acid) motif. However, prelamin A processing into mature lamin A is a unique process because it results in the production of farnesylated and carboxymethylated peptides. In cells from patients with Hutchinson–Gilford progeria syndrome, the mutant prelamin A protein, progerin, cannot release its prenylated carboxyl-terminal moiety and therefore remains permanently associated with the nuclear envelope (NE), causing severe nuclear alterations and a dysmorphic morphology. To obtain a better understanding of the abnormal interaction and retention of progerin in the NE, we analyzed the spatiotemporal distribution of the EGFP fusion proteins with or without a nuclear localization signal (NLS) and a functional CAAX motif in HeLa cells transfected with a series of plasmids that encode the carboxy-terminal ends of progerin and prelamin A. The farnesylated carboxy-terminal fusion peptides bind to the NE and induce the formation of abnormally shaped nuclei. In contrast, the unfarnesylated counterparts exhibit a diffuse localization in the nucleoplasm, without obvious NE deformation. High levels of farnesylated prelamin A and progerin carboxy-terminal peptides induce nucleophagic degradation of the toxic protein, including several nuclear components and chromatin. However, SUN1, a constituent of the linker of nucleoskeleton and cytoskeleton (LINC) complex, is excluded from these autophagic NE protrusions. Thus, nucleophagy requires NE flexibility, as indicated by SUN1 delocalization from the elongated NE–autophagosome complex. Full article

Dictyostelium centrosomes consist of a nucleus-associated cylindrical, three-layered core structure surrounded by a corona consisting of microtubule-nucleation complexes embedded in a scaffold of large coiled-coil proteins. One of them is the conserved CDK5RAP2 protein. Here we focus on the role of Dictyostelium CDK5RAP2 for maintenance of centrosome integrity, its interaction partners and its dynamic behavior during interphase and mitosis. GFP-CDK5RAP2 is present at the centrosome during the entire cell cycle except from a short period during prophase, correlating with the normal dissociation of the corona at this stage. RNAi depletion of CDK5RAP2 results in complete disorganization of centrosomes and microtubules suggesting that CDK5RAP2 is required for organization of the corona and its association to the core structure. This is in line with the observation that overexpressed GFP-CDK5RAP2 elicited supernumerary cytosolic MTOCs. The phenotype of CDK5RAP2 depletion was very reminiscent of that observed upon depletion of CP148, another scaffolding protein of the corona. BioID interaction assays revealed an interaction of CDK5RAP2 not only with the corona markers CP148, γ-tubulin, and CP248, but also with the core components Cep192, CP75, and CP91. Furthermore, protein localization studies in both depletion strains revealed that CP148 and CDK5RAP2 cooperate in corona organization. Full article

Primitive nucleated erythroid cells in the bloodstream have long been suggested to be more similar to nucleated red cells of fish, amphibians, and birds than the red cells of fetal and adult mammals. Rainbow trout Ficoll-purified red blood cells (RBCs) cultured in vitro undergo morphological changes, especially when exposed to stress, and enter a new cell stage that we have coined shape-shifted RBCs (shRBCs). We have characterized these shRBCs using transmission electron microscopy (TEM) micrographs, Wright–Giemsa staining, cell marker immunostaining, and transcriptomic and proteomic evaluation. shRBCs showed reduced density of the cytoplasm, hemoglobin loss, decondensed chromatin in the nucleus, and striking expression of the B lymphocyte molecular marker IgM. In addition, shRBCs shared some features of mammalian primitive pyrenocytes (extruded nucleus surrounded by a thin rim of cytoplasm and phosphatidylserine (PS) exposure on cell surface). These shRBCs were transiently observed in heat-stressed rainbow trout bloodstream for three days. Functional network analysis of combined transcriptomic and proteomic studies resulted in the identification of proteins involved in pathways related to the regulation of cell morphogenesis involved in differentiation, cellular response to stress, and immune system process. In addition, shRBCs increased interleukin 8 (IL8), interleukin 1 β (IL1β), interferon ɣ (IFNɣ), and natural killer enhancing factor (NKEF) protein production in response to viral hemorrhagic septicemia virus (VHSV). In conclusion, shRBCs may represent a novel cell stage that participates in roles related to immune response mediation, homeostasis, and the differentiation and development of blood cells. Full article

Diabetes affects extracellular matrix (ECM) metabolism, contributing to delayed wound healing and lower limb amputation. Application of light (photobiomodulation, PBM) has been shown to improve wound healing. This study aimed to evaluate the influence of PBM on cell adhesion molecules (CAMs) in diabetic wound healing. Isolated human skin fibroblasts were grouped into a diabetic wounded model. A diode laser at 660 nm with a fluence of 5 J/cm² was used for irradiation and cells were analysed 48 h post-irradiation. Controls consisted of sham-irradiated (0 J/cm²) cells. Real-time reverse transcription (RT) quantitative polymerase chain reaction (qPCR) was used to determine the expression of CAM-related genes. Ten genes were up-regulated in diabetic wounded cells, while 25 genes were down-regulated. Genes were related to transmembrane molecules, cell–cell adhesion, and cell–matrix adhesion, and also included genes related to other CAM molecules. PBM at 660 nm modulated gene expression of various CAMs contributing to the increased healing seen in clinical practice. There is a need for new therapies to improve diabetic wound healing. The application of PBM alongside other clinical therapies may be very beneficial in treatment. Full article

The variability in centrosome size, shape, and activity among different organisms provides an opportunity to understand both conserved and specialized actions of this intriguing organelle. Centrosomes in the model organism Dictyostelium sp. share some features with fungal systems and some with vertebrate cell lines and thus provide a particularly useful context to study their dynamics. We discuss two aspects, centrosome positioning in cells and their interactions with nuclei during division as a means to highlight evolutionary modifications to machinery that provide the most basic of cellular services. Full article

Cell–matrix adhesion and cell migration are physiologically important processes that also play a major role in cancer spreading. In cultured cells, matrix adhesion depends on integrin-containing contacts such as focal adhesions. Flotillin-1 and flotillin-2 are frequently overexpressed in cancers and are associated with poor survival. Our previous studies have revealed a role for flotillin-2 in cell–matrix adhesion and in the regulation of the actin cytoskeleton. We here show that flotillins are important for cell migration in a wound healing assay and influence the morphology and dynamics of focal adhesions. Furthermore, anchorage-independent growth in soft agar is enhanced by flotillins. In the absence of flotillins, especially flotillin-2, phosphorylation of focal adhesion kinase and extracellularly regulated kinase is diminished. Flotillins interact with α-actinin, a major regulator of focal adhesion dynamics. These findings are important for understanding the molecular mechanisms of how flotillin overexpression in cancers may affect cell migration and, especially, enhance metastasis formation. Full article

The natural resistance-associated macrophage protein gene (Nramp), has been identified as one of the significant candidate genes responsible for modulating vertebrate natural resistance to intracellular pathogens. Here, we identified and characterized a new Nramp family member, named as maNramp, in the blunt snout bream. The full-length cDNA of maNramp consists of a 153 bp 5′UTR, a 1635 bp open reading frame encoding a protein with 544 amino acids, and a 1359 bp 3′UTR. The deduced protein (maNRAMP) possesses the typical structural features of NRAMP protein family, including 12 transmembrane domains, three N-linked glycosylation sites, and a conserved transport motif. Phylogenetic analysis revealed that maNRAMP shares the significant sequence consistency with other teleosts, and shows the higher sequence similarity to mammalian Nramp2 than Nramp1. It was found that maNramp expressed ubiquitously in all normal tissues tested, with the highest abundance in the spleen, followed by the head kidney and intestine, and less abundance in the muscle, gill, and kidney. After lipopolysaccharide (LPS) stimulation, the mRNA level of maNramp was rapidly up-regulated, which reached a peak level at 6 h. Altogether, these results indicated that maNramp might be related to fish innate immunity and similar to mammalian Nramp1 in function. Full article

Microglia are the resident immune cells in the brain that constitute the brain’s innate immune system. Recent studies have revealed various functions of microglia in the development and maintenance of the central nervous system (CNS) in both health and disease. However, the role of microglia in epilepsy remains largely undiscovered, partly because of the complex phenotypes of activated microglia. Activated microglia likely exert different effects on brain function depending on the phase of epileptogenesis. In this review, we mainly focus on the animal models of temporal lobe epilepsy (TLE) and discuss the proepileptic and antiepileptic roles of activated microglia in the epileptic brain. Specifically, we focus on the roles of microglia in the production of inflammatory cytokines, regulation of neurogenesis, and surveillance of the surrounding environment in epilepsy. Full article

Junctional adhesion molecules (JAMs) are cell surface adhesion receptors of the immunoglobulin superfamily. JAMs are involved in a variety of biological processes both in the adult organism but also during development. These include processes such as inflammation, angiogenesis, hemostasis, or epithelial barrier formation, but also developmental processes such as hematopoiesis, germ cell development, and development of the nervous system. Several of these functions of JAMs depend on a physical and functional interaction with integrins. The JAM – integrin interactions in trans regulate cell-cell adhesion, their interactions in cis regulate signaling processes originating at the cell surface. The JAM – integrin interaction can regulate the function of the JAM as well as the function of the integrin. Beyond the physical interaction with integrins, JAMs can regulate integrin function through intracellular signaling indicating an additional level of JAM – integrin cross-talk. In this review, we describe the various levels of the functional interplay between JAMs and integrins and the role of this interplay during different physiological processes. Full article

The blood–brain barrier (BBB) is a critical biological structure that prevents damage to the brain and maintains its bathing microenvironment. However, this barrier is also the obstacle to deliver beneficial drugs to treat CNS (central nervous system) diseases. Many efforts have been made for improvement of delivering drugs across the BBB in recent years to treat CNS diseases. In this review, the anatomical and functional structure of the BBB is comprehensively discussed. The mechanisms of BBB penetration are summarized, and the methods and effects on increasing BBB permeability are investigated in detail. It also elaborates on the physical, chemical, biological and nanocarrier aspects to improve drug delivery penetration to the brain and introduces some specific drug delivery effects on BBB permeability. Full article