scholarly journals Subcellular Localization of a Novel Alternative Splicing of IIIG9 and Colocalization with PPP1gamma Isoforms

2008 ◽  
Vol 14 (S3) ◽  
pp. 141-143
Author(s):  
C. Sousa ◽  
A.P. Vintém ◽  
M. Fardilha ◽  
O. da Cruz e Silva ◽  
E. da Cruz e Silva
Keyword(s):  
Alternative Splicing ◽  
Male Infertility ◽  
Cell Types ◽  
Sperm Cells ◽  
Regulatory Subunits ◽  
Specific Antibodies ◽  
Fluorescent Microscope ◽  
Bovine Sperm ◽  
Spermatogonia Cells ◽  
Different Cell Types

In testis we find mainly PPP1gamma2 isoform. We hypothesize that in different cell types we can find different regulatory subunits that may constitute targets for therapeutics of diseases such as male infertility, cancer and Alzheimer's disease. We identified a novel alternative splicing isoform of IIIG9 in testis, a known regulator of PPP1, IIIG9sT, and the aim of this study was its further characterization. We used a specific antibody for IIIG9sT in order to characterize its localization in bovine sperm cells. We also transfected IIIG9sT-GFP construct in mouse spermatogonia cells (GC-1 cells) and we used specific antibodies for each PPP1 isoform for the colocalization studies. We observed them under a fluorescent microscope and a LSM and quantified a high co-localization with PPP1gamma1 and 2 isoforms.

scholarly journals Assembly of multiple dystrobrevin-containing complexes in the kidney

2000 ◽  
Vol 113 (15) ◽  
pp. 2715-2724
Author(s):  
N.Y. Loh ◽  
S.E. Newey ◽  
K.E. Davies ◽  
D.J. Blake
Keyword(s):  
Skeletal Muscle ◽  
Protein Complexes ◽  
Cell Types ◽  
Epithelial Morphogenesis ◽  
Basal Region ◽  
Molecular Architecture ◽  
Renal Epithelial Cells ◽  
Specific Antibodies ◽  
Different Cell Types ◽  
Deficient Mice

Dystrophin is the key component in the assembly and maintenance of the dystrophin-associated protein complex (DPC) in skeletal muscle. In kidney, dystroglycan, an integral component of the DPC, is involved in kidney epithelial morphogenesis, suggesting that the DPC is important in linking the extracellular matrix to the internal cytoskeleton of kidney epithelia. Here, we have investigated the molecular architecture of dystrophin-like protein complexes in kidneys from normal and dystrophin-deficient mice. Using isoform-specific antibodies, we show that the different cell types that make up the kidney maintain different dystrophin-like complexes. These complexes can be broadly grouped according to their dystrobrevin content: beta-dystrobrevin containing complexes are present at the basal region of renal epithelial cells, whilst alpha-dystrobrevin-1 containing complexes are found in endothelial and smooth muscle cells. Furthermore, these complexes are maintained even in the absence of all dystrophin isoforms. Thus our data suggest that the functions and assembly of the dystrophin-like complexes in kidney differ from those in skeletal muscle and implicate a protein other than dystrophin as the primary molecule in the assembly and maintenance of kidney complexes. Our findings also provide a possible explanation for the lack of kidney pathology in Duchenne muscular dystrophy patients and mice lacking all dystrophin isoforms.

scholarly journals Chromogranin A in the Human Gastrointestinal Tract: An Immunocytochemical Study with Region-specific Antibodies

2002 ◽  
Vol 50 (11) ◽  
pp. 1487-1492 ◽  
Author(s):  
Guida Maria Portela-Gomes ◽  
Mats Stridsberg
Keyword(s):  
Chromogranin A ◽  
Cell Types ◽  
The Other ◽  
Gi Tract ◽  
Human Gastrointestinal Tract ◽  
Double Immunofluorescence ◽  
Gastrin Cells ◽  
Specific Antibodies ◽  
Ec Cells ◽  
Different Cell Types

We studied the immunoreactivity of 12 different region-specific antibodies to the chromogranin A (CgA) molecule in the various neuroendocrine cell types of the human gastrointestinal (GI) tract by using double immunofluorescence techniques. These staining results were compared with others obtained with a commercial monoclonal CgA antibody (LK2H10). G (gastrin)-cells showed immunoreactivity to virtually all region-specific antibodies, but with varying frequency. Most intestinal EC (enterochromaffin)- and L (enteroglucagon)-cells were immunoreactive to the antibodies to the N-terminal and mid-portion of the CgA molecule, whereas the EC-cells in the stomach reacted with fewer region-specific antibodies. D (somatostatin)-cells reacted to the CgA 411–424 antibody and only occasionally showed immunoreactivity to the other CgA antibodies. A larger cytoplasmic area was stained with the antibodies to CgA 17–38 and 176–195 than with the other antibodies tested. These differences in staining pattern may reflect different cleavage of the CgA molecule in different cell types and at different regions of the GI tract.

scholarly journals Combinatorial Control of Exon Recognition

2007 ◽  
Vol 283 (3) ◽  
pp. 1211-1215 ◽  
Author(s):  
Klemens J. Hertel
Keyword(s):  
Alternative Splicing ◽  
Splice Site ◽  
Cell Types ◽  
Splice Sites ◽  
Splice Site Selection ◽  
Multiple Parameters ◽  
Splicing Regulators ◽  
Eukaryotic Genes ◽  
Different Cell Types ◽  
Mature Mrnas

Pre-mRNA splicing is a fundamental process required for the expression of most metazoan genes. It is carried out by the spliceosome, which catalyzes the removal of noncoding intronic sequences to assemble exons into mature mRNAs prior to export and translation. Given the complexity of higher eukaryotic genes and the relatively low level of splice site conservation, the precision of the splicing machinery in recognizing and pairing splice sites is impressive. Introns ranging in size from <100 up to 100,000 bases are removed efficiently. At the same time, a large number of alternative splicing events are observed between different cell types, during development, or during other biological processes. This extensive alternative splicing implies a significant flexibility of the spliceosome to identify and process exons within a given pre-mRNA. To reach this flexibility, splice site selection in higher eukaryotes has evolved to depend on multiple parameters such as splice site strength, the presence or absence of splicing regulators, RNA secondary structures, the exon/intron architecture, and the process of pre-mRNA synthesis itself. The relative contributions of each of these parameters control how efficiently splice sites are recognized and flanking introns are removed.

scholarly journals Alternative Splicing Modulates Autoinhibition and SH3 Accessibility in the Src Kinase Fyn

2009 ◽  
Vol 29 (24) ◽  
pp. 6438-6448 ◽  
Author(s):  
C. Brignatz ◽  
M. P. Paronetto ◽  
S. Opi ◽  
M. Cappellari ◽  
S. Audebert ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Src Kinase ◽  
Cell Types ◽  
Site Directed Mutagenesis ◽  
Fine Tuning ◽  
Control Mechanisms ◽  
Molecular Control ◽  
Fyn Kinase ◽  
Functional Consequences ◽  
Different Cell Types

ABSTRACT Src family kinases are central regulators of a large number of signaling pathways. To adapt to the idiosyncrasies of different cell types, these kinases may need a fine-tuning of their intrinsic molecular control mechanisms. Here, we describe on a molecular level how the Fyn kinase uses alternative splicing to adapt to different cellular environments. Using structural analysis, site-directed mutagenesis, and functional analysis, we show how the inclusion of either exon 7A or 7B affects the autoinhibition of Fyn and how this changes the SH3-dependent interaction and tyrosine phosphorylation of Sam68, with functional consequences for the Sam68-regulated survival of epithelial cells. Our results illustrate a novel mechanism of evolution that may contribute to the complexity of Src kinase regulation.

scholarly journals MicroRNAs: Recent insights towards their role in male infertility and reproductive cancers

2019 ◽  
Vol 19 (1) ◽  
pp. 31-42 ◽  
Author(s):  
Muhammad Babar Khawar ◽  
Rabia Mehmood ◽  
Nabila Roohi
Keyword(s):  
Male Infertility ◽  
Reproductive System ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Regulatory Mechanisms ◽  
Aberrant Expression ◽  
Protein Coding ◽  
Reproductive Cancers ◽  
Post Transcriptional Regulation ◽  
Different Cell Types

Spermatogenesis is a tightly controlled, multi-step process in which mature spermatozoa are produced. Disruption of regulatory mechanisms in spermatogenesis can lead to male infertility, various diseases of male reproductive system, or even cancer. The spermatogenic impairment in infertile men can be associated with different etiologies, and the exact molecular mechanisms are yet to be determined. MicroRNAs (miRNAs) are a type of non-protein coding RNAs, about 22 nucleotides long, with an essential role in post-transcriptional regulation. miRNAs have been recognized as important regulators of various biological processes, including spermatogenesis. The aim of this review is to summarize the recent literature on the role of miRNAs in spermatogenesis, male infertility and reproductive cancers, and to evaluate their potential in diagnosis, prognosis and therapy of disease. Experimental evidence shows that aberrant expression of miRNAs affects spermatogenesis at multiple stages and in different cell types, most often resulting in infertility. In more severe cases, dysregulation of miRNAs leads to cancer. miRNAs have enormous potential to be used as diagnostic and prognostic markers as well as therapeutic targets in male infertility and reproductive system diseases. However, to exploit this potential fully, we need a better understanding of miRNA-mediated regulation of spermatogenesis, including the characterization of yet unidentified miRNAs and related regulatory mechanisms.

scholarly journals MALDI-IMS combined with shotgun proteomics identify and localize new factors in male infertility

2021 ◽  
Vol 4 (3) ◽  
pp. e202000672
Author(s):  
Shibojyoti Lahiri ◽  
Wasim Aftab ◽  
Lena Walenta ◽  
Leena Strauss ◽  
Matti Poutanen ◽  
...  
Keyword(s):  
Male Infertility ◽  
Imaging Mass Spectrometry ◽  
Shotgun Proteomics ◽  
Cell Types ◽  
The Novel ◽  
Proteomic Approach ◽  
Maldi Ims ◽  
Tissue Proteomics ◽  
Different Cell Types

Spermatogenesis is a complex multi-step process involving intricate interactions between different cell types in the male testis. Disruption of these interactions results in infertility. Combination of shotgun tissue proteomics with MALDI imaging mass spectrometry is markedly potent in revealing topological maps of molecular processes within tissues. Here, we use a combinatorial approach on a characterized mouse model of hormone induced male infertility to uncover misregulated pathways. Comparative testicular proteome of wild-type and mice overexpressing human P450 aromatase (AROM+) with pathologically increased estrogen levels unravels gross dysregulation of spermatogenesis and emergence of pro-inflammatory pathways in AROM+ testis. In situ MS allowed us to localize misregulated proteins/peptides to defined regions within the testis. Results suggest that infertility is associated with substantial loss of proteomic heterogeneity, which define distinct stages of seminiferous tubuli in healthy animals. Importantly, considerable loss of mitochondrial factors, proteins associated with late stages of spermatogenesis and steroidogenic factors characterize AROM+ mice. Thus, the novel proteomic approach pinpoints in unprecedented ways the disruption of normal processes in testis and provides a signature for male infertility.

scholarly journals PSXIII-5 The age dynamics of the spermatogenic cells development in the roosters’ testes

2019 ◽  
Vol 97 (Supplement_3) ◽  
pp. 373-373
Author(s):  
Anastasia N Vetokh ◽  
Natalia A Volkova ◽  
Evgeniya K Tomgorova ◽  
Ludmila A Volkova ◽  
Natalia A Zinovieva
Keyword(s):  
Germ Cells ◽  
Sertoli Cells ◽  
Genetic Material ◽  
Cell Types ◽  
Seminiferous Tubules ◽  
Sperm Cells ◽  
Spermatogenic Cells ◽  
Different Cell Types ◽  
Hematoxylin Eosin ◽  
Testis Tissue

Abstract The cells of the male gonads are considered as a valuable genetic material for the conservation of the gene pool of breeds and lines of agricultural birds, as well as the directed modification of the poultry genome. Mature germ cells – spermatozoa and their predecessors – spermatogonia, spermatocytes and spermatids can be used for these purposes. To obtain these types of cells, it is necessary to know the characteristics of their development (spermatogenesis). The dynamics of the development of certain spermatogenic cell types in the testicular tubules of different-aged roosters has been studied. Histological studies were performed on testes of roosters aged from 1 week to 6 months with an interval of 2 weeks. Samples of testis tissue were fixed in Bouin’s solution. Histological sections were stained with hematoxylin-eosin. Identification of different cell types (Sertoli, spermatogonia, spermatocytes, spermatids, sperm cells) was carried out according to their morphology. At the age of 1–6 weeks in the seminiferous tubule of roosters, the mainly presence of two cell types was noted: Sertoli cells and spermatogonia. From 7 weeks of age, spermatocytes were detected in the seminiferous tubules, in the 4 months - spermatids, in the 5.5 months - sperm cells. The number of Sertoli cells remained almost unchanged with age and was 21 ± 2. The percentage of these cells decreased with age from 71 ± 3 % to 5 ± 1 %. The percentage of spermatogonia also decreased with age from 75 ± 2 % to 7 ± 1 %. The number of spermatids and spermatozoa, on the contrary, increased to puberty (6 months) and reached 54 %. The study was supported by the RFBR within Project no.18-29-07079.

scholarly journals Alternative splicing tunes sodium channels to support channel- and neuron-specific effects

2020 ◽  
Author(s):  
Gabriele Lignani ◽  
Andrianos Liavas ◽  
Dimitri M Kullmann ◽  
Stephanie Schorge
Keyword(s):  
Alternative Splicing ◽  
Action Potential ◽  
Sodium Channels ◽  
Neuronal Excitability ◽  
Cell Types ◽  
Alternate Splicing ◽  
Specific Effects ◽  
Voltage Gated Sodium Channels ◽  
Temporal Encoding ◽  
Different Cell Types

AbstractNeuronal excitability is tightly regulated, requiring rapidly activating and inactivating voltage-gated sodium channels to allow accurate temporal encoding of information. Alternative splicing greatly broadens the repertoire of channels, but the adaptive significance of this phenomenon is incompletely understood. An alternative splicing event that is conserved across vertebrates affects part of the first domain of sodium channels and modulates their availability after inactivation. Here we use this conserved splicing event to ask whether this modulation has consistent effects in different neuronal backgrounds, or whether a conserved splicing event can be exploited to produce distinct effects in different cell types. We show that the consequences of alternate splicing of human Nav1.1 and Nav1.2 for neuronal activity depend on whether they are expressed in the cell types where they normally predominate (interneurons or excitatory neurons, respectively). Splicing in the ‘adult’ isoform in both channels is sufficient to slow action potential rise times in all neurons. However, changes to both action potential half width and maximal firing rate are specific to cell type and channel, with each channel appearing tuned to mediate effects in its predominant neuronal background. Finally, we use dynamic clamp to demonstrate that alternative splicing in Nav1.1 changes how interneurons fire during epileptiform events. Our data show that, for sodium channels, despite conserved amino acid changes and similar effects on channel gating, alternative splicing has distinct impacts on neuronal properties, thus highlighting how closely sodium channels are tuned to distinct cellular backgrounds.

Gephyrin: does splicing affect its function?

2006 ◽  
Vol 34 (1) ◽  
pp. 45-47 ◽  
Author(s):  
I. Paarmann ◽  
T. Saiyed ◽  
B. Schmitt ◽  
H. Betz
Keyword(s):  
Alternative Splicing ◽  
Ion Channels ◽  
Single Cells ◽  
Protein Structures ◽  
Cell Types ◽  
Binding Partners ◽  
Its Gene ◽  
Substantial Progress ◽  
Different Cell Types ◽  
Made In

Gephyrin is a protein involved in both synaptic anchoring of inhibitory ligand-gated ion channels and molybdenum cofactor synthesis. Substantial progress has been made in understanding its gene and protein structures. Furthermore, numerous binding partners of gephyrin have been identified. The mechanisms by which these interactions occur are unclear at present. Alternative splicing has been proposed to contribute to gephyrin's functional diversity within single cells as well as in different cell types and tissues.

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