Guimarães Alessander O, Motta Fabiana L, Alves Viviane S, Castilho Beatriz A, Pesquero João B
Departamento de Biofísica, Universidade Federal de São Paulo, São Paulo, Brazil.
BMC Mol Biol. 2009 Feb 8;10:7. doi: 10.1186/1471-2199-10-7.
A major effort of the scientific community has been to obtain complete pictures of the genomes of many organisms. This has been accomplished mainly by annotation of structural and functional elements in the genome sequence, a process that has been centred in the gene concept and, as a consequence, biased toward protein coding sequences. Recently, the explosion of transcriptome data generated and the discovery of many functional non-protein coding RNAs have painted a more detailed and complex scenario for the genome. Here we analyzed the mouse carboxypeptidase M locus in this broader perspective in order to define the mouse CPM gene structure and evaluate the existence of other transcripts from the same genomic region.
Bioinformatic analysis of nucleotide sequences that map to the mouse CPM locus suggests that, in addition to the mouse CPM mRNA, it expresses at least 33 different transcripts, many of which seem to be non-coding RNAs. We randomly chose to evaluate experimentally four of these extra transcripts. They are expressed in a tissue specific manner, indicating that they are not artefacts or transcriptional noise. Furthermore, one of these four extra transcripts shows expression patterns that differed considerably from the other ones and from the mouse CPM gene, suggesting that there may be more than one transcriptional unit in this locus. In addition, we have confirmed the mouse CPM gene RefSeq sequence by rapid amplification of cDNA ends (RACE) and directional cloning.
This study supports the recent view that the majority of the genome is transcribed and that many of the resulting transcripts seem to be non-coding RNAs from introns of genes or from independent transcriptional units. Although some of the information on the transcriptome of many organisms may actually be artefacts or transcriptional noise, we argue that it can be experimentally evaluated and used to find and define biological functional elements on the genome. Furthermore, the transcription of other functional RNAs besides the protein coding RNA from a specific genomic locus imposes extra care when designing and interpreting experiments involving genetic manipulations or expression detection and quantification.
科学界的一项主要工作是获取许多生物体基因组的完整图谱。这主要通过对基因组序列中的结构和功能元件进行注释来完成，该过程一直以基因概念为核心，因此偏向于蛋白质编码序列。最近，转录组数据的激增以及许多功能性非蛋白质编码RNA的发现，为基因组描绘了一幅更详细、更复杂的图景。在这里，我们从这个更广阔的视角分析了小鼠羧肽酶M基因座，以确定小鼠CPM基因结构，并评估同一基因组区域中其他转录本的存在情况。
对定位到小鼠CPM基因座的核苷酸序列进行生物信息学分析表明，除了小鼠CPM mRNA外，它还表达至少33种不同的转录本，其中许多似乎是非编码RNA。我们随机选择对其中四种额外的转录本进行实验评估。它们以组织特异性方式表达，表明它们不是人为产物或转录噪声。此外，这四种额外转录本中的一种显示出与其他转录本以及小鼠CPM基因有很大不同的表达模式，这表明该基因座可能存在不止一个转录单元。此外，我们通过cDNA末端快速扩增（RACE）和定向克隆证实了小鼠CPM基因的RefSeq序列。
本研究支持了最近的观点，即基因组的大部分都被转录，并且许多产生的转录本似乎是来自基因内含子或独立转录单元的非编码RNA。尽管关于许多生物体转录组的一些信息实际上可能是人为产物或转录噪声，但我们认为可以通过实验进行评估，并用于在基因组上发现和定义生物功能元件。此外，在设计和解释涉及基因操作或表达检测与定量的实验时，除了来自特定基因组位点的蛋白质编码RNA外，其他功能性RNA的转录需要格外小心。
