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Regional and developmental brain expression patterns of SNAP25 splice variants.

Prescott GR, Chamberlain LH - BMC Neurosci (2011)

Bottom Line: Differential splicing of the SNAP25 gene results in the expression of two transcripts, SNAP25a and SNAP25b.The extent of this up-regulation in SNAP25b expression was similar across cortex, cerebellum and hippocampus.The antibodies generated and characterized in this study represent important tools for future analyses of these essential SNARE protein isoforms.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Integrative Physiology, School of Biomedical Sciences, Hugh Robson Building, University of Edinburgh, Edinburgh EH8 9XD, UK.

ABSTRACT

Background: SNAP25 is an essential SNARE protein for regulated exocytosis in neuronal cells. Differential splicing of the SNAP25 gene results in the expression of two transcripts, SNAP25a and SNAP25b. These splice variants differ by only 9 amino acids, and studies of their expression to date have been limited to analysis of the corresponding mRNAs. Although these studies have been highly informative, it is possible that factors such as differential turnover of the SNAP25 proteins could complicate interpretations based entirely on mRNA expression profiles.

Results: We report the generation and characterization of antibodies that distinguish between SNAP25a and SNAP25b isoforms, and their use to investigate the expression profile of these proteins in rat and human brain. In rat brain, SNAP25b protein expression increased dramatically during post-natal development, whereas the increase in SNAP25a expression was more modest and variable. The extent of this up-regulation in SNAP25b expression was similar across cortex, cerebellum and hippocampus. The SNAP25 isoforms also displayed distinct regional expression patterns, with SNAP25a very weakly expressed in both rat and human cerebellum. Quantitative analysis revealed that SNAP25b was the dominant isoform in all adult human brain regions examined.

Conclusions: SNAP25a and SNAP25b display distinct developmental and regional expression profiles in rat and human brain. These differences might reflect distinct functions of these highly conserved isoforms in membrane fusion pathways in the brain. The antibodies generated and characterized in this study represent important tools for future analyses of these essential SNARE protein isoforms.

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Related in: MedlinePlus

Regional expression of SNAP25a and SNAP25b in rat brain. Lysates from various brain regions (supplied by Zyagen; 3 animals/sample; ~ 10 week old) were resolved by SDS-PAGE and transferred to nitrocellulose for immunoblotting analysis using antibodies against SNAP25a (3067), SNAP25b (3068), panSNAP25, SNAP23 or actin. Position of molecular weight markers are shown on the left side of all blots.
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Figure 3: Regional expression of SNAP25a and SNAP25b in rat brain. Lysates from various brain regions (supplied by Zyagen; 3 animals/sample; ~ 10 week old) were resolved by SDS-PAGE and transferred to nitrocellulose for immunoblotting analysis using antibodies against SNAP25a (3067), SNAP25b (3068), panSNAP25, SNAP23 or actin. Position of molecular weight markers are shown on the left side of all blots.

Mentions: To compare the regional expression levels of SNAP25a and SNAP25b in brain, samples of hippocampus, hypothalamus, thalamus, cerebellum and cortex from ~10 week-old rats were analyzed by immunoblotting. SNAP25a displayed highest levels (per μg of total protein) in cortex and hypothalamus (Figure 3), whereas SNAP25b had the highest immunoreactivity in cortex and cerebellum lysates. A marked difference in regional expression levels between the SNAP25 isoforms was detected in cerebellum: SNAP25b expression was highest in this brain region, whereas SNAP25a expression was lowest (Figure 3). Probing the brain region lysates with a panSNAP25 antibody revealed that total SNAP25 expression levels mirrored those of SNAP25b (Figure 3, third panel), consistent with the idea that SNAP25b is the major SNAP25 isoform expressed in adult brain.


Regional and developmental brain expression patterns of SNAP25 splice variants.

Prescott GR, Chamberlain LH - BMC Neurosci (2011)

Regional expression of SNAP25a and SNAP25b in rat brain. Lysates from various brain regions (supplied by Zyagen; 3 animals/sample; ~ 10 week old) were resolved by SDS-PAGE and transferred to nitrocellulose for immunoblotting analysis using antibodies against SNAP25a (3067), SNAP25b (3068), panSNAP25, SNAP23 or actin. Position of molecular weight markers are shown on the left side of all blots.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3104942&req=5

Figure 3: Regional expression of SNAP25a and SNAP25b in rat brain. Lysates from various brain regions (supplied by Zyagen; 3 animals/sample; ~ 10 week old) were resolved by SDS-PAGE and transferred to nitrocellulose for immunoblotting analysis using antibodies against SNAP25a (3067), SNAP25b (3068), panSNAP25, SNAP23 or actin. Position of molecular weight markers are shown on the left side of all blots.
Mentions: To compare the regional expression levels of SNAP25a and SNAP25b in brain, samples of hippocampus, hypothalamus, thalamus, cerebellum and cortex from ~10 week-old rats were analyzed by immunoblotting. SNAP25a displayed highest levels (per μg of total protein) in cortex and hypothalamus (Figure 3), whereas SNAP25b had the highest immunoreactivity in cortex and cerebellum lysates. A marked difference in regional expression levels between the SNAP25 isoforms was detected in cerebellum: SNAP25b expression was highest in this brain region, whereas SNAP25a expression was lowest (Figure 3). Probing the brain region lysates with a panSNAP25 antibody revealed that total SNAP25 expression levels mirrored those of SNAP25b (Figure 3, third panel), consistent with the idea that SNAP25b is the major SNAP25 isoform expressed in adult brain.

Bottom Line: Differential splicing of the SNAP25 gene results in the expression of two transcripts, SNAP25a and SNAP25b.The extent of this up-regulation in SNAP25b expression was similar across cortex, cerebellum and hippocampus.The antibodies generated and characterized in this study represent important tools for future analyses of these essential SNARE protein isoforms.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Integrative Physiology, School of Biomedical Sciences, Hugh Robson Building, University of Edinburgh, Edinburgh EH8 9XD, UK.

ABSTRACT

Background: SNAP25 is an essential SNARE protein for regulated exocytosis in neuronal cells. Differential splicing of the SNAP25 gene results in the expression of two transcripts, SNAP25a and SNAP25b. These splice variants differ by only 9 amino acids, and studies of their expression to date have been limited to analysis of the corresponding mRNAs. Although these studies have been highly informative, it is possible that factors such as differential turnover of the SNAP25 proteins could complicate interpretations based entirely on mRNA expression profiles.

Results: We report the generation and characterization of antibodies that distinguish between SNAP25a and SNAP25b isoforms, and their use to investigate the expression profile of these proteins in rat and human brain. In rat brain, SNAP25b protein expression increased dramatically during post-natal development, whereas the increase in SNAP25a expression was more modest and variable. The extent of this up-regulation in SNAP25b expression was similar across cortex, cerebellum and hippocampus. The SNAP25 isoforms also displayed distinct regional expression patterns, with SNAP25a very weakly expressed in both rat and human cerebellum. Quantitative analysis revealed that SNAP25b was the dominant isoform in all adult human brain regions examined.

Conclusions: SNAP25a and SNAP25b display distinct developmental and regional expression profiles in rat and human brain. These differences might reflect distinct functions of these highly conserved isoforms in membrane fusion pathways in the brain. The antibodies generated and characterized in this study represent important tools for future analyses of these essential SNARE protein isoforms.

Show MeSH
Related in: MedlinePlus