PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-6 (6)
 

Clipboard (0)
None

Select a Filter Below

Journals
Authors
more »
Year of Publication
1.  Essential roles of zebrafish rtn4/Nogo paralogues in embryonic development 
Neural Development  2014;9:8.
Background
As a consequence of gene/genome duplication, the RTN4/Nogo gene has two counterparts in zebrafish: rtn4a and rtn4b. The shared presence of four specific amino acid motifs—M1 to M4—in the N-terminal region of mammalian RTN4, and zebrafish Rtn4b suggests that Rtn4b is the closest homologue of mammalian Nogo-A.
Results
To explore their combined roles in zebrafish development, we characterized the expression patterns of rtn4a and rtn4b in a comparative manner and performed morpholino-mediated knockdowns. Although both genes were coexpressed in the neural tube and developing brain at early stages, they progressively acquired distinct expression domains such as the spinal cord (rtn4b) and somites (rtn4a). Downregulation of rtn4a and rtn4b caused severe brain abnormalities, with rtn4b knockdown severely affecting the spinal cord and leading to immobility. In addition, the retinotectal projection was severely affected in both morphants, as the retina and optic tectum appeared smaller and only few retinal axons reached the abnormally reduced tectal neuropil. The neuronal defects were more persistent in rtn4b morphants. Moreover, the latter often lacked pectoral fins and lower jaws and had malformed branchial arches. Notably, these defects led to larval death in rtn4b, but not in rtn4a morphants.
Conclusions
In contrast to mammalian Nogo-A, its zebrafish homologues, rtn4a and particularly rtn4b, are essential for embryonic development and patterning of the nervous system.
doi:10.1186/1749-8104-9-8
PMCID: PMC4113184  PMID: 24755266
Brain and spinal cord development; Larval motility; Morpholino knockdown; Nogo; Reticulon; rtn4; Zebrafish
3.  Acetate Activation in Methanosaeta thermophila: Characterization of the Key Enzymes Pyrophosphatase and Acetyl-CoA Synthetase 
Archaea  2012;2012:315153.
The thermophilic methanogen Methanosaeta thermophila uses acetate as sole substrate for methanogenesis. It was proposed that the acetate activation reaction that is needed to feed acetate into the methanogenic pathway requires the hydrolysis of two ATP, whereas the acetate activation reaction in Methanosarcina sp. is known to require only one ATP. As these organisms live at the thermodynamic limit that sustains life, the acetate activation reaction in Mt. thermophila seems too costly and was thus reevaluated. It was found that of the putative acetate activation enzymes one gene encoding an AMP-forming acetyl-CoA synthetase was highly expressed. The corresponding enzyme was purified and characterized in detail. It catalyzed the ATP-dependent formation of acetyl-CoA, AMP, and pyrophosphate (PPi) and was only moderately inhibited by PPi. The breakdown of PPi was performed by a soluble pyrophosphatase. This enzyme was also purified and characterized. The pyrophosphatase hydrolyzed the major part of PPi (KM = 0.27 ± 0.05 mM) that was produced in the acetate activation reaction. Activity was not inhibited by nucleotides or PPi. However, it cannot be excluded that other PPi-dependent enzymes take advantage of the remaining PPi and contribute to the energy balance of the cell.
doi:10.1155/2012/315153
PMCID: PMC3426162  PMID: 22927778
4.  A Novel Inducible Protein Production System and Neomycin Resistance as Selection Marker for Methanosarcina mazei 
Archaea  2012;2012:973743.
Methanosarcina mazei is one of the model organisms for the methanogenic order Methanosarcinales whose metabolism has been studied in detail. However, the genetic toolbox is still limited. This study was aimed at widening the scope of utilizable methods in this group of organisms. (i) Proteins specific to methanogens are oftentimes difficult to produce in E. coli. However, a protein production system is not available for methanogens. Here we present an inducible system to produce Strep-tagged proteins in Ms. mazei. The promoter p1687, which directs the transcription of methyl transferases that demethylate methylamines, was cloned into plasmid pWM321 and its activity was determined by monitoring β-glucuronidase production. The promoter was inactive during growth on methanol but was rapidly activated when trimethylamine was added to the medium. The gene encoding the β-glucuronidase from E. coli was fused to a Strep-tag and was cloned downstream of the p1687 promoter. The protein was overproduced in Ms. mazei and was purified in an active form by affinity chromatography. (ii) Puromycin is currently the only antibiotic used as a selectable marker in Ms. mazei and its relatives. We established neomycin resistance as a second selectable marker by designing a plasmid that confers neomycin resistance in Ms. mazei.
doi:10.1155/2012/973743
PMCID: PMC3407599  PMID: 22851906
5.  Membrane-Bound Electron Transport in Methanosaeta thermophila▿ 
Journal of Bacteriology  2011;193(11):2868-2870.
The obligate aceticlastic methanogen Methanosaeta thermophila uses a membrane-bound ferredoxin:heterodisulfide oxidoreductase system for energy conservation. We propose that the system is composed of a truncated form of the F420H2 dehydrogenase, methanophenazine, and the heterodisulfide reductase. Hence, the electron transport chain is distinct from those of well-studied Methanosarcina species.
doi:10.1128/JB.00162-11
PMCID: PMC3133127  PMID: 21478356
6.  Function of Ech Hydrogenase in Ferredoxin-Dependent, Membrane-Bound Electron Transport in Methanosarcina mazei▿  
Journal of Bacteriology  2009;192(3):674-678.
Reduced ferredoxin is an intermediate in the methylotrophic and aceticlastic pathway of methanogenesis and donates electrons to membrane-integral proteins, which transfer electrons to the heterodisulfide reductase. A ferredoxin interaction has been observed previously for the Ech hydrogenase. Here we present a detailed analysis of a Methanosarcina mazei Δech mutant which shows decreased ferredoxin-dependent membrane-bound electron transport activity, a lower growth rate, and faster substrate consumption. Evidence is presented that a second protein whose identity is unknown oxidizes reduced ferredoxin, indicating an involvement in methanogenesis from methylated C1 compounds.
doi:10.1128/JB.01307-09
PMCID: PMC2812462  PMID: 19948802

Results 1-6 (6)