mRNA affects protein fate

The genetic code of proteins may dictate much more than their amino acid sequences, a new linkurl:paper;http://www.sciencemag.org/cgi/content/abstract/sci;329/5998/1534?maxtoshow=&hits=10&RESULTFORMAT=&fulltext=Differential+arginylation+of+actin+isoforms+is+regulated+by+coding+sequence-dependent+degradation&searchid=1&FIRSTINDEX=0&resourcetype=HWCIT from __Science__ suggests -- it may hold their ultimate fate.

β-actin Image:Wikimedia commons

linkurl:Anna Kashina;http://www.med.upenn.edu/apps/faculty/index.php/g20000546/p4638831 and her colleagues at the University of Pennsylvania described a novel mechanism of protein regulation whereby differences in the mRNA sequences of two nearly identical proteins cause them to be translated into proteins at different speeds. As a result, one form is targeted for quick degradation while the other goes on to carry out important functions in the cell. "It's an innovative piece of work," said Chloë linkurl:Bulinski,;http://www.columbia.edu/cu/biology/faculty-data/chloe-bulinski/faculty.html a cell biologist at Columbia University who was not involved in the study. "It's an example in which two proteins that have different nucleotide sequences but almost the same amino acid sequence can behave very differently." For many years, scientists had been puzzled by the two versions, or isoforms, of the structural protein actin, known as β and γ. Although they are encoded by different genes, their amino acid sequences differ by only four amino acids (out of a total of nearly 400), and they behave very similarly when isolated in test tubes. "They seem to be able to substitute for each other," Kashina said. But in living cells -- specifically non-muscle cells -- they play strikingly different roles. β-actin is mostly found near one edge of the cell, where it forms the filaments that push the membrane forward and help cells move. γ-actin, on the other hand, forms actin structures deeper inside cells, such as stress fibers that compose the cytoskeleton and give the cell its shape. "Nobody understood the difference in function because they are nearly identical," Kashina said. However, a breakthrough came in 2006 when Kashina and her colleagues found that a posttranslational modification known as arginylation, in which the amino acid arginine is attached to the N-terminus of the protein, was critical for directing β-actin to the edges of cells. But the researchers still scratched their heads over why β-actin was arginylated and shuttled off to the cell's edge while its nearly identical twin was not. "We suspected that the difference was in their stability," Kashina said. Indeed, when they compared levels of artificially arginylated β and γ-actin in cells, they found that arginylated γ-actin was being degraded at a much faster rate. "Both isoforms can be arginylated," she added, "but one of them [γ-actin] is just immediately degraded so we never find it." At the amino acid level, there is no difference that explains why arginylation selectively dooms γ-actin and not β. Therefore, the researchers turned to the nucleotide sequences of their mRNAs, which happen to differ substantially. They changed the four codons in the mRNA encoding γ-actin so that the resulting amino acid sequence produced is the same as β-actin. Despite the fact that it was now identical to β-actin at the protein level, it was still as unstable as normal γ-actin. The reverse was also true. Using β-actin's mRNA to code for γ-actin, increased the levels of the naturally unstable isoform. Furthermore, β-actin accumulated much more quickly in cells that lacked the protein degradation machinery, suggesting that in addition to being degraded more slowly than γ-actin, it was being synthesized faster. These results suggested that there was something at the mRNA level -- something in the silent nucleotides -- that was causing the functional differences between the two forms of actin. "The RNA that's encoding γ, but not the one that's encoding β, is able to get sort of tangled up," cell biologist Chloë Bulinski explained. "It's able to form a structure that can't be unfurled and therefore this slows down the rate of translation." Slower translation, it turned out, increased γ-actin's likelihood of being degraded by exposing a particular lysine amino acid -- a prime target for ubiquination, one of the cell's protein degradation pathways. Once the protein is complete, the lysine is hidden deep inside, "but while the protein is synthesized this lysine is exposed," Kashina explained. Because the ubiquitination machinery is attracted to the arginine addition, the arginylated actins are more likely to be degraded than their unmanipulated counterparts. But because β-actin's translation is quicker, it buries its lysine before the ubiquitin marks it for destruction. Arginylated γ-actin, on the other hand, is not fast enough, and only the un-arginylated form tends to survive. The result is arginylated β-actin, which is shuttled to the cell's edge, and normal γ-actin, which contributes to the cell's cytoskeleton. "Never before had people realized that differences in coding sequence can lead to different regulation of proteins," Kashina said. "The new notion is that posttranslational modifications are in a way encoded into the coding sequence of a protein, and not just occur based on the amino acid sequence after the protein is finished." F. Zhang, et al., "Differential arginylation of actin isoforms is regulated by coding sequence-dependent degradation," Science, 329:1534-37, 2010.
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[16th April 2001]*linkurl:Tagged for Cleansing;http://www.the-scientist.com/article/display/55707/
[June 2009]*linkurl:Protein Degradation;http://www.the-scientist.com/article/display/17482/
[31st March 1997]