scholarly journals De novo formation of basal bodies in Naegleria gruberi

2005 ◽  
Vol 169 (5) ◽  
pp. 719-724 ◽  
Author(s):  
Hong-Kyung Kim ◽  
Jeong-Gu Kang ◽  
Shigehiko Yumura ◽  
Charles J. Walsh ◽  
Jin Won Cho ◽  
...  
Keyword(s):  
Western Blotting ◽  
Basal Body ◽  
Gel Electrophoresis ◽  
De Novo ◽  
Myosin Ii ◽  
Basal Bodies ◽  
2D Gel Electrophoresis ◽  
2D Gel ◽  
Naegleria Gruberi

The de novo formation of basal bodies in Naegleria gruberi was preceded by the transient formation of a microtubule (MT)-nucleating complex containing γ-tubulin, pericentrin, and myosin II complex (GPM complex). The MT-nucleating activity of GPM complexes was maximal just before the formation of visible basal bodies and then rapidly decreased. The regulation of MT-nucleating activity of GPM complexes was accomplished by a transient phosphorylation of the complex. Inhibition of dephosphorylation after the formation of basal bodies resulted in the formation of multiple flagella. 2D-gel electrophoresis and Western blotting showed a parallel relationship between the MT-nucleating activity of GPM complexes and the presence of hyperphosphorylated γ-tubulin in the complexes. These data suggest that the nucleation of MTs by GPM complexes precedes the de novo formation of basal bodies and that the regulation of MT-nucleating activity of GPM complexes is essential to the regulation of basal body number.

scholarly journals Naegleria gruberi De Novo Basal Body Assembly Occurs via Stepwise Incorporation of Conserved Proteins

2010 ◽  
Vol 9 (6) ◽  
pp. 860-865 ◽  
Author(s):  
Lillian K. Fritz-Laylin ◽  
Zoe June Assaf ◽  
Sean Chen ◽  
W. Zacheus Cande
Keyword(s):  
Basal Body ◽  
De Novo ◽  
Unicellular Eukaryote ◽  
Basal Bodies ◽  
Microtubule Cytoskeleton ◽  
Cytoplasmic Microtubule ◽  
Content Type ◽  
Genes Encoding ◽  
Associated Proteins ◽  
Naegleria Gruberi

ABSTRACT Centrioles and basal bodies are discrete structures composed of a cylinder of nine microtubule triplets and associated proteins. Metazoan centrioles can be found at mitotic spindle poles and are called basal bodies when used to organize microtubules to form the core structure of flagella. Naegleria gruberi, a unicellular eukaryote, grows as an amoeba that lacks a cytoplasmic microtubule cytoskeleton. When stressed, Naegleria rapidly (and synchronously) differentiates into a flagellate, forming a complete cytoplasmic cytoskeleton de novo, including two basal bodies and flagella. Here, we show that Naegleria has genes encoding conserved centriole proteins. Using novel antibodies, we describe the localization of three centrosomal protein homologs (SAS-6, γ-tubulin, and centrin-1) during the assembly of the flagellate microtubule cytoskeleton. We also used these antibodies to show that Naegleria expresses the proteins in the same order as their incorporation into basal bodies, with SAS-6 localizing first, followed by centrin and finally γ-tubulin. The similarities between basal body assembly in Naegleria and centriole assembly in animals indicate that mechanisms of assembly, as well as structure, have been conserved throughout eukaryotic evolution.

Identification of nuclei associated proteins by 2D-gel electrophoresis and mass spectrometry

2001 ◽  
Vol 109 (1) ◽  
pp. 3-11 ◽  
Author(s):  
Jonas Bergquist ◽  
Johan Gobom ◽  
Anders Blomberg ◽  
Peter Roepstorff ◽  
Rolf Ekman
Keyword(s):  
Mass Spectrometry ◽  
Gel Electrophoresis ◽  
2D Gel Electrophoresis ◽  
Associated Proteins ◽  
2D Gel

scholarly journals Differential methods of cell lysis for protein profiling of Alexandrium sp. using 2D gel electrophoresis

2020 ◽  
Vol 15 (1) ◽  
Author(s):  
Nurul Ashima Hamdan ◽  
Normawaty Mohammad-Nor ◽  
Shafida Abd. Hamid ◽  
Nor Hasniza Md. Zin ◽  
Noraslinda Muhamad Bunnori
Keyword(s):  
Gel Electrophoresis ◽  
Cold Water ◽  
Protein Extraction ◽  
Excellent Result ◽  
Axenic Culture ◽  
Cell Lysis ◽  
Protein Profiling ◽  
2D Gel Electrophoresis ◽  
Fundamental Study ◽  
2D Gel

Introduction: Protein profiling of harmful algae is an ongoing study where the latest analysis was conducted on A. minutum. It is a fundamental study where the protein expression of targeted species can be used to understand the biochemical pathway of selected proteins. The Malaysia Alexandrium spp. has the potential to cause massive blooming that brings harm to the aquatic ecosystem. Methods: In this experiment three methods of cell lysis were tested against A. leei isolated from Malaysian waters. Results: The Axenic culture of the sample was established in enriched seawater media (ESDK) with 12 hours of light and 12 hours of dark conditions. The sample was extracted during exponential phase (day 18) where the same amount of cells was collected via centrifugation. The same buffer was used for each technique of cell lysis in order to get the best protein profiles in terms of a band or spot intensity and number. The cells of A. leei were lysed using sonication in the ice-cold water bath, sonication with probe and freeze-thawing followed by sonication in iced bath is the best method of protein extraction. Conclusions: Nevertheless, it can be concluded that the qualitative analysis using SDS-PAGE and 2D gel electrophoresis could be best method and the freeze thawing mode of cell lysis produced an excellent result among others as the protein spots produced were precise and less streaking were observed.

Sign in / Sign up

Export Citation Format

Share Document