scholarly journals CEP290 tethers flagellar transition zone microtubules to the membrane and regulates flagellar protein content

2010 ◽  
Vol 190 (5) ◽  
pp. 927-940 ◽  
Author(s):  
Branch Craige ◽  
Che-Chia Tsao ◽  
Dennis R. Diener ◽  
Yuqing Hou ◽  
Karl-Ferdinand Lechtreck ◽  
...  
Keyword(s):  
Protein Content ◽  
Transition Zone ◽  
Biochemical Analysis ◽  
Close Association ◽  
Protein Composition ◽  
Intraflagellar Transport ◽  
Transport Proteins ◽  
Flagellar Membrane ◽  
Perinatal Lethality ◽  
Flagellar Protein

Mutations in human CEP290 cause cilia-related disorders that range in severity from isolated blindness to perinatal lethality. Here, we describe a Chlamydomonas reinhardtii mutant in which most of the CEP290 gene is deleted. Immunoelectron microscopy indicated that CEP290 is located in the flagellar transition zone in close association with the prominent microtubule–membrane links there. Ultrastructural analysis revealed defects in these microtubule–membrane connectors, resulting in loss of attachment of the flagellar membrane to the transition zone microtubules. Biochemical analysis of isolated flagella revealed that the mutant flagella have abnormal protein content, including abnormal levels of intraflagellar transport proteins and proteins associated with ciliopathies. Experiments with dikaryons showed that CEP290 at the transition zone is dynamic and undergoes rapid turnover. The results indicate that CEP290 is required to form microtubule–membrane linkers that tether the flagellar membrane to the transition zone microtubules, and is essential for controlling flagellar protein composition.

scholarly journals WD60/FAP163 is a dynein intermediate chain required for retrograde intraflagellar transport in cilia

2013 ◽  
Vol 24 (17) ◽  
pp. 2668-2677 ◽  
Author(s):  
Ramila S. Patel-King ◽  
Renée M. Gilberti ◽  
Erik F. Y. Hom ◽  
Stephen M. King
Keyword(s):  
Functional Role ◽  
Biochemical Analysis ◽  
Intraflagellar Transport ◽  
Dynein Light Chain ◽  
Orthologous Protein ◽  
Dynein Intermediate Chain ◽  
Flagellar Protein ◽  
Assembly Defect ◽  
Intermediate Chain

Retrograde intraflagellar transport (IFT) is required for assembly of cilia. We identify a Chlamydomonas flagellar protein (flagellar-associated protein 163 [FAP163]) as being closely related to the D1bIC(FAP133) intermediate chain (IC) of the dynein that powers this movement. Biochemical analysis revealed that FAP163 is present in the flagellar matrix and is actively trafficked by IFT. Furthermore, FAP163 copurified with D1bIC(FAP133) and the LC8 dynein light chain, indicating that it is an integral component of the retrograde IFT dynein. To assess the functional role of FAP163, we generated an RNA interference knockdown of the orthologous protein (WD60) in planaria. The Smed-wd60(RNAi) animals had a severe ciliary assembly defect that dramatically compromised whole-organism motility. Most cilia were present as short stubs that had accumulated large quantities of IFT particle–like material between the doublet microtubules and the membrane. The few remaining approximately full-length cilia had a chaotic beat with a frequency reduced from 24 to ∼10 Hz. Thus WD60/FAP163 is a dynein IC that is absolutely required for retrograde IFT and ciliary assembly.

Proteomics-Based Characterization of the Effects of MMP14 on the Protein Content of Exosomes from Corneal Fibroblasts

2020 ◽  
Vol 27 (10) ◽  
pp. 979-988
Author(s):  
Kyu-Yeon Han ◽  
Jin-Hong Chang ◽  
Dimitri T. Azar
Keyword(s):  
Protein Content ◽  
Catalytic Domain ◽  
Protein Composition ◽  
Proteomics Analysis ◽  
Knock Out ◽  
Corneal Fibroblast ◽  
Flow Cytometric ◽  
Corneal Fibroblasts ◽  
Liquid Chromatography Tandem Mass

Background: Exosomes secreted by corneal fibroblasts contain matrix metalloproteinase (MMP) 14, which is known to influence pro-MMP2 accumulation on exosomes. Accordingly, we hypothesized that the enzymatic activity of MMP14 may alter the protein content of corneal fibroblast- secreted exosomes. Objective: The aim of this study was to investigate the effects of MMP14 on the composition and biological activity of corneal fibroblast-derived exosomes. Methods: Knock out of the catalytic domain (ΔExon4) of MMP14 in corneal fibroblasts was used to determine the effect of MMP14 expression on the characteristics of fibroblast-secreted exosomes. The amount of secreted proteins and their size distribution were measured using Nano Tracking Analysis. Proteins within exosomes from wild-type (WT) and ΔExon4-deficient fibroblasts were identified by liquid chromatography-tandem mass spectrometry (MS/MS) proteomics analysis. The proteolytic effects of MMP14 were evaluated in vitro via MS identification of eliminated proteins. The biological functions of MMP14-carrying exosomes were investigated via fusion to endothelial cells and flow cytometric assays. Results: Exosomes isolated from WT and ΔExon4-deficient fibroblasts exhibited similar size distributions and morphologies, although WT fibroblasts secreted a greater amount of exosomes. The protein content, however, was higher in ΔExon4-deficient fibroblast-derived exosomes than in WT fibroblast-derived exosomes. Proteomics analysis revealed that WT-derived exosomes included proteins that regulated cell migration, and ΔExon4 fibroblast-derived exosomes contained additional proteins that were cleaved by MMP14. Conclusion: Our findings suggest that MMP14 expression influences the protein composition of exosomes secreted by corneal fibroblasts, and through those biological components, MMP14 in corneal fibroblasts derived-exosomes may regulate corneal angiogenesis.

Hedgehog signalling in the mouse requires intraflagellar transport proteins

Nature ◽  
2003 ◽  
Vol 426 (6962) ◽  
pp. 83-87 ◽  
Author(s):  
Danwei Huangfu ◽  
Aimin Liu ◽  
Andrew S. Rakeman ◽  
Noel S. Murcia ◽  
Lee Niswander ◽  
...  
Keyword(s):  
Intraflagellar Transport ◽  
Transport Proteins ◽  
Hedgehog Signalling

scholarly journals The Effect of Abiotic Stresses on the Protein Composition of Four Hungarian Wheat Varieties

Plants ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 1
Author(s):  
Dalma Nagy-Réder ◽  
Zsófia Birinyi ◽  
Marianna Rakszegi ◽  
Ferenc Békés ◽  
Gyöngyvér Gell
Keyword(s):  
Abiotic Stress ◽  
Protein Content ◽  
Functional Properties ◽  
Crop Production ◽  
Global Climate ◽  
Protein Composition ◽  
Grain Protein Content ◽  
Wheat Varieties ◽  
Significant Difference ◽  
Negative Effect

Global climate change in recent years has resulted in extreme heat and drought events that significantly influence crop production and endanger food security. Such abiotic stress during the growing season has a negative effect on yield as well as on the functional properties of wheat grain protein content and composition. This reduces the value of grain, as these factors significantly reduce end-use quality. In this study, four Hungarian bread wheat cultivars (Triticum aestivum ssp. aestivum) with different drought and heat tolerance were examined. Changes in the size- and hydrophobicity-based distribution of the total proteins of the samples have been monitored by SE- and RP-HPLC, respectively, together with parallel investigations of changes in the amounts of the R5 and G12 antibodies related to celiac disease immunoreactive peptides. Significant difference in yield, protein content and composition have been observed in each cultivar, altering the amounts of CD-related gliadin, as well as the protein parameters directly related to techno-functional properties (Glu/Gli ratio, UPP%). The extent of changes largely depended on the timing of the abiotic stress. The severity of the negative effect depended on the growth stage in which abiotic stress occurred.

Quality traits of wheat determined by small-scale dough testing methods

2001 ◽  
Vol 52 (12) ◽  
pp. 1325 ◽  
Author(s):  
F. Békés ◽  
P. W. Gras ◽  
R. S. Anderssen ◽  
R. Appels
Keyword(s):  
Protein Content ◽  
Molecular Genetic ◽  
Protein Composition ◽  
Quality Parameters ◽  
Molecular Genetic Analysis ◽  
Small Scale ◽  
Quality Traits ◽  
Testing Procedures ◽  
Large Populations ◽  
Wheat Lines

The dough properties of flours from the grain of 172 doubled haploid lines of a Cranbrook Halberd cross, grown at 3 locations, were determined with traditional and small-scale dough testing equipment. The experiments were aimed at determining the genetic factors that underpin the flour processing properties of wheat flour. Seven mixing parameters determined on a 2-g Mixograph™, as well as the maximum resistance (RMAX) and extensibility (EXT) measured on a Micro-Extension Tester, were identified as quality traits for genetic mapping studies, to identify the underlying quantitative trait loci (QTL). For each of the 3 locations in which the wheat lines were grown, relationships between the quality parameters and genetic markers were constructed for the populations. The associations of HMW- and LMW-glutenin allele combinations with the quality traits were investigated using ANOVA, linear parametric, and non-parametric methods. Of particular interest were qualitative and quantitative assessments of the extremes of the quality traits in each population. The relative contributions of the glutenincoding loci to quality were determined and it was found that the growing conditions to which wheat lines were subjected significantly affected the analyses. The nature and extent of these variations could not be explained by changes in protein content alone, and were related to environmentally induced alterations in the protein composition. From a comparison of the measurements made with the small-scale Mixograph™ with those from both the Extensograph™ and a Micro-Extension Tester, it was concluded that the same information about RMAX and EXT obtained from traditional extension testing could be obtained using small-scale dough tests. The data provided a direct validation for the application of small-scale testing for the screening of large populations. The comparisons of large and small scale testing procedures also provided the basis defining a new trait, ‘M-extensibility’, which is obtained from protein content and selected Mixograph data. This parameter was able to be measured more accurately and was shown to be closely related to the traditional extensibility measurement, and thus very useful for molecular/genetic analysis. The M-extensibility trait could be mapped as a major QTL to LMW-glutenin subunit loci on chromosomes 1B and 1D.

scholarly journals Variation in the Protein Composition of Human Milk during Extended Lactation: A Narrative Review

Nutrients ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 1124 ◽  
Author(s):  
Sergio Verd ◽  
Gemma Ginovart ◽  
Javier Calvo ◽  
Jaume Ponce-Taylor ◽  
Antoni Gaya
Keyword(s):  
Breast Milk ◽  
Human Milk ◽  
Protein Content ◽  
Milk Protein ◽  
Mammalian Species ◽  
Protein Composition ◽  
Developed Countries ◽  
First Year ◽  
Second Year ◽  
First Year Of Life

The aim of this review is to evaluate changes in protein parameters in the second year postpartum. There is considerable agreement among authors about the declining trend of human milk protein concentrations, but most research on protein content in breast milk focuses on the first year of life and comes from developed countries. Whereas this is the case for exclusive breastfeeding or for breastfeeding into the first year of life, the opposite applies to weaning or extended breastfeeding. This review is predominantly based on observational epidemiological evidence and on comparative research linking breast milk composition with cutting down on breastfeeding. Studies dating back several decades have shown an increase in the proportion of immunoglobulins, lactoferrin, and serum albumin during weaning. According to the limited data available, it seems likely that the regulation of milk protein composition during involution can be ascribed to alterations in tight junctions. In studies on humans and other mammalian species, offspring suckle more from mothers that produce more dilute milk and the increase in milk protein concentration is positively correlated to a decrease in suckling frequency during weaning. High milk protein contents were first reported in nonindustrial communities where breastfeeding is sustained the longest, but recent papers from urbanized communities have taken credit for rediscovering the increase in protein content of human milk that becomes evident with prolonged breastfeeding. This review presents an overview of the changes in breast milk protein parameters in the second year postpartum to enable milk banks’ practitioners to make informed nutritional decisions on preterm infants.

scholarly journals FURTHER STUDIES ON THE CHEMISTRY OF BLOOD SERUM

1913 ◽  
Vol 17 (4) ◽  
pp. 444-452 ◽  
Author(s):  
Albert A. Epstein
Keyword(s):  
Blood Serum ◽  
Protein Content ◽  
Protein Composition ◽  
The Other ◽  
Total Protein Content ◽  
Protein Nitrogen ◽  
Standard Values ◽  
The Individual ◽  
Chronic Interstitial Nephritis ◽  
Functional Deficiency

The following conclusions may be drawn from the results obtained in this study. 1. In the minor surgical cases (considered normal) the chemical composition of the blood serum agrees, as far as its proteins are concerned, with the usual standard values. When examined at different times, the serum of such cases shows no variation in the total protein content or in its individual fractions. The incoagulable nitrogen, however, varies considerably in the total amount in the different cases, as well as in its percentage relations to the other constituents of the serum. 2. The cases of prostatic hypertrophy, with or without chronic interstitial nephritis, show no change from the normal in the character of the protein composition of the serum, nor in the ratio which the individual fractions bear to each other. On the other hand, the incoagulable and non-protein nitrogen of the serum show marked fluctuations, some of which correspond to the degree of functional deficiency of the kidneys. 3. In cases of localized infections of the kidneys, the changes in the blood serum are twofold. One concerns the proteins, and is traceable to the infection; and the other concerns the non-protein nitrogen, results from functional impairment, and varies with the amount of destruction of the kidney substance. Thus an increase in the globulins is observed in these sera similar to that occurring in infections in other localities; the non-protein content increases apparently in direct proportion to the degree of deficiency of the kidney, and becomes diminished when the function of the kidneys improves.

scholarly journals Centrioles initiate cilia assembly but are dispensable for maturation and maintenance in C. elegans

2017 ◽  
Vol 216 (6) ◽  
pp. 1659-1671 ◽  
Author(s):  
Daniel Serwas ◽  
Tiffany Y. Su ◽  
Max Roessler ◽  
Shaohe Wang ◽  
Alexander Dammermann
Keyword(s):  
Fluorescence Microscopy ◽  
Transition Zone ◽  
Electron Tomography ◽  
Intraflagellar Transport ◽  
Basal Bodies ◽  
C Elegans ◽  
Cilia Formation ◽  
Syndrome Protein

Cilia are cellular projections that assemble on centriole-derived basal bodies. While cilia assembly is absolutely dependent on centrioles, it is not known to what extent they contribute to downstream events. The nematode C. elegans provides a unique opportunity to address this question, as centrioles do not persist at the base of mature cilia. Using fluorescence microscopy and electron tomography, we find that centrioles degenerate early during ciliogenesis. The transition zone and axoneme are not completely formed at this time, indicating that cilia maturation does not depend on intact centrioles. The hydrolethalus syndrome protein HYLS-1 is the only centriolar protein known to remain at the base of mature cilia and is required for intraflagellar transport trafficking. Surprisingly, targeted degradation of HYLS-1 after initiation of ciliogenesis does not affect ciliary structures. Taken together, our results indicate that while centrioles are essential to initiate cilia formation, they are dispensable for cilia maturation and maintenance.

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