Researchers have uncovered a new class of small, non-coding RNAs in worms, according to a report in Cell
. Using massively parallel sequencing technology, David Bartel
, a Howard Hughes Medical Institute Investigator at the Massachusetts Institute of Technology, and colleagues sequenced some 400,000 small RNAs from Caenorhabditis elegans
, identifying 18 new microRNA genes and more than 5,000 RNAs (called 21U-RNAs) whose common features are a 5' uridine residue, a shared upstream motif and chromosomal location, and a length of precisely 21 nucleotides.
There is "no question" of the significance of these findings, according to Phillip Sharp
, Institute Professor at MIT, who is collaborating with Bartel to understand the biology of 21U-RNAs but was not involved in the current study. "We made the fundamental mistake over the last 30 years of not paying attention to non-coding RNAs," Sharp told The Scientist
. "RNA interference and microRNAs tell us that small RNAs, 20 nucleotides or so long, have more than enough information in them to give you very specific gene regulation."
"This discovery further emphasizes the point that we have not completely plumbed the depths of RNA function, and we are excited to see how this develops," Scott Baskerville, a research scientist at the RNA biotechnology firm Dharmacon
and a former Bartel lab postdoc, told The Scientist
in an email. "It will be interesting to see if using a similar strategy (deep sequencing) new classes of small RNAs are discovered in other species," said Baskerville, who was not involved in the current study.
Bartel's group has been systematically working its way through the RNA census of C. elegans
for several years now. In 2001, the group sequenced
330 small RNAs using traditional Sanger sequencing to identify 55 microRNAs. Bartel followed that project up in 2003 with a deeper sequencing effort
, reading 4,000 small RNAs to yield an additional 40 or so microRNAs. In this latest study, his team used a massively parallel pyrosequencing system from 454 Life Sciences
to read some 400,000 small RNAs, including 18 new microRNAs.
"We know that there must be more microRNAs that we haven't yet sequenced, but we don't know how many more," Bartel said. "It does seem like we're reaching diminishing returns." One of the microRNAs he knows to be missing is lsy-6
, which has been identified genetically but never actually sequenced. Thought to be expressed in only one to nine cells in the worm, and small cells at that, lsy-6
could be 100,000 times less abundant than a microRNA expressed in all cells of the worm, Bartel estimated.
Also identified in the sequencing data were several thousand endogenous, small interfering RNAs, plus 5,700 or so 21U-RNAs, all of which are the same length, start with a 5'-uridine, terminate with a modified 3' ribose, and share an upstream DNA sequence motif. Representing about nine percent of all the sequence reads, the 21U-RNAs mostly mapped to two discrete regions of chromosome IV.
"What's striking," said Sharp, is that "there's a lot of specificity in the biochemical description of these RNAs that suggest they reflect something that's almost certainly important in the organism."
Using the upstream sequence motif as a guide, Bartel estimates there are 12,000 or so candidate 21U-RNA loci in the C. elegans
genome. "If you think of each locus as a gene, then you've almost doubled the number of genes in the worm," he said. Surprisingly, though the genomic location of these genes is conserved between C. elegans
and the related C. briggsae
, the sequence of the transcript itself is not. "It almost seems like evolution is maximizing the diversity of these small RNAs, rather than maximizing their conservation," Bartel said.
One open question is the function of 21U-RNAs. Given the lack of sequence conservation between C. elegans
and C. briggsae
, Bartel speculated they could serve to affect local chromatin structure and gene expression by affecting the distribution of histone proteins, for instance. Other unanswered questions include which polymerase synthesizes 21U-RNAs and how are they processed. Bartel said he is working with 2006 Nobel laureate Craig Mello
of the University of Massachusetts Medical Center to determine whether mutant worms that are defective in RNAi show defective 21U-RNA production, too.
Said Sharp, "I'd be surprised if we don't within a year or so have some idea what they might be doing."
Jeffrey M. Perkel
Links within this article:
J.G. Ruby et al., "Large-scale sequencing reveals 21U-RNAs and additional microRNAs and endogenous siRNAs in C. elegans," Cell, 127:1193 - 1207, Dec. 15, 2006.
N.C. Lau et al., "An abundant class of tiny RNAs with probable regulatory roles in Caenorhabditis elegans
, 294:858 - 62, 2001.
L.P. Lim et al., "The microRNAs of Caenorhabditis elegans
," Genes Dev
, 17:991 - 1008, 2003.
454 Life Sciences
K. Grens, "Fire and Mello win Nobel Prize," The Scientist
, Oct. 2, 2006