On the function of lincRNAs

There's plenty of evidence that most of the DNA in mammalian genomes is junk [Five Things You Should Know if You Want to Participate in the Junk DNA Debate]. There's also plenty of evidence that as much as 10% of these genomes are functional in some way or another. This is a lot more DNA than the amount in coding regions but that shouldn't surprise anyone since we've known about functional noncoding DNA for half a century.

Lot's of genes specify functional RNA molecules. The best known ones are the genes for ribosomal RNAs, tRNAs, the spliceosomal RNAs, and a variety of other catalytic RNAs. A host of small regulatory RNAs have been characterized in bacteria over the past five decades (Waters and Storz, 2009) and in the past few decades a variety of different types of small RNAs have been identified in eukaryotes (see Sharp, 2009). These include miRNAs, siRNAs, piRNAs, and others (Malone and Hannon, 2009; Carthew and Sontheimer, 2009).

Theme

Genomes
& Junk DNASome of the most interesting RNAs are the long intergenic noncoding or lincRNAs (Ponting et al. 2009). The average size of these longer RNAs is about 1500 bp and there are about 1600 conserved lincRNA genes in the mammalian genome (Guttman et al., 2009). This makes up only about 0.08% of the genome but these RNAs are very curious.

Mammalian genomes are pervasively transcribed. Most of the transcripts are present at less than one copy per cell and they are not very stable. Their sequences are not conserved. Thus, they have all the characteristics of spurious transcripts and are undoubtedly junk RNA (Struhl, 2007). That's not what we're talking about here although lots of people are confused.

What we're talking about is true functional RNAs transcribed from noncoding DNA. They've been around for a long time and many are well characterized. We want to understand what the others are doing.

One way to decide if the genes for these RNAs are actually doing something is to disrupt them by knocking them out and looking for an effect. That's what Savageau et al. (2013) did with 18 genes for mouse link RNAs. They found that five of the mutant strains of mice had severe developmental defects that were often lethal (Fendrr, Peril, and Mdgt: mice with a deleted Mdgt lincRNA gene are shown in the photo). Two other strains, linc-Brn1b, and linc-Pint had less severe developmental defects.

What this shows it that five out of eighteen lincRNA genes are very important in mouse development. Some of the others may also be functional but the phenotypes may not as obvious. We now know functions for several dozen lincRNA genes. It is still an open question whether there are over a thousand functional lincRNA genes in mammalian genomes or whether most of them are spurious transcripts. Keep in mind that even if every single one is functional, it makes no significant impact on the amount of junk DNA in the genome.

What I like about this paper (Savageau et al., 2013) is that the authors are aware of the controversy. Here's what they say in the Discussion ...

In the post genomic era, thousands of long noncoding RNAs have been discovered as transcribed units in mammalian genomes. However, what fraction of these new transcripts have general functional significance in vivo is debated. While several studies have indicated a role for lincRNAs in diverse biological processes (Ponting et al., 2009; Rinn and Chang, 2012; Mercer and Mattick, 2013; Ulitsky and Bartel, 2013), it has been suggested that most transcripts could represent nonfunctional transcriptional by-products (Struhl, 2007; Kowalczyk et al., 2012). Early critical studies of knockout strains (e.g., Xist and Tsix) did find lncRNAs implicated in X inactivation to be required for life. Yet, of the relatively few lncRNA mouse models derived since, many have displayed subtle defects or no phenotype (Ripoche et al., 1997; Gordon et al., 2010; Anguera et al., 2011; Nakagawa et al., 2011; Zhang et al., 2012).

It's actually quite rare for people working on these RNAs to admit that most of them might not have a function.

John Mattick is a fierce opponent of junk DNA. He's never met an RNA that he didn't think was functional and he seemed in the past to be completely unaware of the evidence for junk DNA. Mattick wrote a review of the Savageau et al. et al. paper because he's quite excited about the fact that some more lincRNAs seem to be functional. Unfortunately, his review gets off on the wrong foot by stating that functional noncoding RNAs were only discovered in the 1990s and by perpetuating his false story about the correlation between genome size and complexity. Here's the beginning of his article ...

It has been known since the late 1970s that many DNA sequences are transcribed but not translated. Moreover, most protein-coding genes in mammals are fragmented, with only a small fraction of the primary RNA transcript being spliced together to form messenger RNA. For many years it was assumed that untranslated RNA molecules served no useful purpose but, starting in the mid-1990s, a small body of researchers, including the present author (Mattick, 1994), have been arguing that these RNAs transmit regulatory information, possibly associated with the emergence of multicellular organisms. This is supported by the observation that the proportion of noncoding genomic sequences broadly correlates with developmental complexity, reaching over 98% in mammals (Liu et al., 2013), although others have argued that the increase in genome size is due to the inefficiency of selection against non-functional elements as body size goes up and population size goes down (Lynch, 2007).

The good news is that Mattick seems more willing that usual to admit to the controversy about the function of these small RNAs.

Because many lncRNAs appear to be expressed at low levels, and many have lower sequence conservation than messenger RNAs, one interpretation has been that these RNAs represent transcriptional noise from complex genomes cluttered with evolutionary debris. However, assessments of sequence conservation rely on assumptions about the non-functionality and representative distribution of reference sequences, which are not verified and cannot be directly tested (Pheasant and Mattick, 2007). Nonetheless, many lncRNAs show patches of relative sequence conservation (Derrien et al., 2012), and even more do so at the secondary structural level (Smith et al., 2013).

Mattick still thinks that most of the lincRNA genes are doing something important. He falls back on the fact that a large number of these genes are expressed in mammalian brain tissue and it may be difficult to detect a phenotype in mice if the cognitive defects are subtle.

Whatever.

The point is that scientists are beginning to find out what some of these lincRNAs are doing but that it has nothing to do with the junk DNA debate.

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Malone, C.D. and Hannon, G.J. (2009) Small RNAs as guardians of the genome. Cell 136, 656-668. [doi: 10.1016/j.cell.2009.01.045]

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