Actin is a structural protein found in all eukaryotic cells, where it performs the important function of supporting cell structure. In human cells, there are three types of actin present: alpha, beta and gamma.
Alpha actins are found mostly in muscle (smooth and cardiac) and skeletal cells, whereas other cells tend to have beta and gamma actin. Filaments of actin mechanically support cell shape and also, through their motion, can be used by cells for transport within the cell and cellular movement.
The motion of actin filaments is also used to position organelles such as the Golgi apparatus, mitochondria, and peroxisomes within cells, to adjust cell polarity and to regulate junctions between cells.1
The behavior and activity of actin can become extremely complicated and is regulated by interactions with other factors and signals present within the cell.2
The Role of Actin in the Cytoplasm and Nucleus
Actin found in the cytoplasm is polymerized via an ATP-promoted mechanism into long, twisted filaments of high strength and stability. The start of the polymerization occurs slowly with the aggregation of actin-ATP complexes into short oligomers of low stability, which can readily break down.
However, once an oligomer reaches a critical size, they become stable enough to act as a point of nucleation; resulting in rapid filament growth at both ends of the filament as more actin-ATP complexes aggregate.
Following aggregation, the ATP bound to the actin is cleaved by hydrolysis to give ADP. As this only occurs during elongation of the chain the ends of the filament are capped with ATP. Research has shown that ATP hydrolysis is not essential for actin formation as polymerization can still occur when using non-hydrolysable ATP analogs.3
There are more than 100 different proteins that, which regulate small variations in the actin filament and its activity. Regulation can take the form of influencing assembly and production rates, reducing filament length and also encouraging cross-link formation to produce actin bundles.2,4
Actin has also been shown to operate within the nucleus of cells. Historically the presence of actin within the nucleus was understood to be an artifact. However, research has demonstrated that actin plays roles in the regulation of transcription and the remodeling of chromatin.5
To illustrate, nuclear actin is a key factor in multiple chromatin-altering machines, where it attaches to chromatin modifiers (e.g. NuA4 HAT) or remodelers (e.g. INO80, SWI-SNF and SWR1).6 Actin is also believed to regulate the initiation of transcription via interactions with RNA polymerases (I, II and III7) and with pre-mRNA.
Using Actin as a Loading Control
Actin isoforms have a very high level of similarity to one another with only small differences in their primary amino acid sequence. For example, when comparing beta cyto-actin and gamma-cyto actin only four amino acid residues are different. This indicates that preservation of actin structures is under a high level of evolutionary pressure.1
Information about the homology of actin types and across different species is required for the selection of antibodies and interpretation of the bands in western blots.
Conventionally, beta actin has been used as the loading control in life science research as it has been assumed that the level of protein present is unaffected by the cell types composition, stage of development or treatment. However, recent research has begun to question if this assumption is correct.
An example of this is the use of beta-actin in experiments. This is now advised against because beta-actin expression has been found to vary depending on how mature a tissue is, this is especially significant for research involving tissues of different types or maturities.8,9
References and Further Reading
- Alberts B, Johnson A, Lewis J, et al. (2002). Molecular Biology of the Cell. 4th edition.
- Budnar S, Yap AS (2013). A mechanobiological perspective on cadherins and the actin-myosin cytoskeleton. F1000Prime Rep. Sep 2; 5:35.
- Lodish H, Berk A, Zipursky SL et al (2000). Molecular Cell Biology. 4th edition. New York: W. H. Freeman.
- Mullins RD, Hansen SD (2013). In vitro studies of actin filament and network dynamics. Curr Opin Cell Biol. Feb;25(1):6-13. New York: Garland Science.
- de Lanerolle P (2012). Nuclear actin and myosins at a glance. J Cell Sci. November 1; 125(21): 4945–4949.
- Szerlong H, Hinata K, Viswanathan R, Erdjument-Bromage H, Tempst P, Cairns BR (2008).The HSA domain binds nuclear actin-related proteins to regulate chromatin remodeling ATPases. Nat Struct Mol Biol.15(5):469-76.
- Percipalle P (2013). Co-transcriptional nuclear actin dynamics. Nucleus. Jan-Feb;4(1):43-52.
- Dittmer A, Dittmer J (2006). Beta-actin is not a reliable loading control in Western blot analysis. Electrophoresis. Jul; 27(14):2844-5.
- Eaton SL, Roche SL, Llavero Hurtado M, Oldknow KJ, Farquharson C, Gillingwater TH, Wishart TM (2013). Total protein analysis as a reliable loading control for quantitative fluorescent Western blotting. PLoS One. Aug 30;8(8):e72457.
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