On 2014 Oct 18, Pavel Baranov commented:

What is the difference between Open Reading Frame (ORF) and Coding Sequence (CDS)?

Thank you for the reply. I think the disagreement lies in our understanding of what Open Reading Frame is.

A simple and effective definition of ORF is a sequence of codons not interrupted with stop codons: nucleotide sequence is open for reading in one of the three (for RNA) or six (dsRNA) frames. ORF is an abstract notion, it can be found in any sequence. Both protein-codng and non-coding sequences have ORFs.

Coding Sequence (CDS) is the part of RNA that encodes protein. CDS often, but not always (exceptions are ribosomal frameshifting, stop codon readthrough, etc.), is located within a single ORF. A single ORF may contain more than one Coding Region if, for example, translation begins at different start codons within the same ORF.

From your reply I presume that you suggest to define ORF as a sequence of codons from start to stop (like CDS). But the problem with this detention is that it is unclear what should we consider as a start of ORF. AUG? But not all AUGs are starts and not all starts are AUGs. Also there are no starts in non-coding RNAs, but there are ORFs in non-coding sequences.

Besides if we use this definition, all eukaryotic mRNAs coding for multiple protein isoforms would need to be described as bi- or even polycistronic.

I hope that this discussion brings some clarity to the terminology used or at least it draws attention to a potential confusion when terms such as ORF, CDS and cistron are not explicitly defined.

On 2014 Oct 03, Jonathan C Kagan commented:

In response to the comment by Baranov, we would first like to thank you for your interest in our work. We agree that alternatively translated eukaryotic proteins most commonly share a reading frame, as our study mentioned. In fact, we discuss this point in the analysis of our ribosomal profiling data. For instance, we observed that truncations are more common than internal out-of-frame translation products. Regardless of whether the products share a reading frame, however, a point of interest is the regulation allowing ribosomes to initiate translation at more than one location on a transcript. We chose to use the terms polycistronic and bicistronic for two reasons. First, bicistronic mRNAs are operationally defined as transcripts that produce two stable proteins of distinct functions, regardless of whether the two proteins share sequence similarity. Our study clearly demonstrated that this is the case with the MAVS transcript. Whether these functionally distinct proteins share coding sequence with each other is functionally irrelevant. Second, a transcript producing a truncated protein (such as MAVS) conforms to generally accepted definitions of a polycistronic transcript, such as you provided: “…protein products of distinct coding ORFs are translated from the same transcript.” While the two MAVS proteins are encoded in the same reading frame, their production is initiated at unique start sites; thus, they have distinct coding sequences. In short, we consider them to be distinct ORFs that share a reading frame and therefore produce two proteins from a bicistronic mRNA.

On 2014 Aug 06, Pavel Baranov commented:

What is a polycistronic mRNA?

This is an interesting research article that provides insights into distinct functions of two proteoforms that differ at their N-termini due to the use of alternative translation initiation starts. Terming the corresponding mRNA bicistronic is, however, somewhat misleading. When we refer to a bacterial mRNA as polycistronic we imply that protein products of distinct coding ORFs are translated from the same transcript. Here the ORF is the same, but the translation initiates at different codons of that ORF. There are several examples of human proteins with truncated or extended N-termini produced from alternative translation initiation starts, see a recently discovered extension of PTEN for a startling example, see Hopkins BD, 2013. True bicistronic mRNAs are rare. Molybdopterin subunits MOCS2A and MOCS2B were reported to be produced from two different but overlapping ORFs at the same mRNA, see Stallmeyer B, 1999. If this were true the corresponding mRNA could be classified as bicistronic, however, further evidence suggested that these proteins are produced from alternatively spliced transcript variants, see Hahnewald R, 2006. A good example of an eukaryotic polycistronic mRNA could be found in Drosophilla tal (aka pri) mRNA that codes for four peptides produced from four distinct ORFs, see Kondo T, 2010. Perhaps, human mRNAs with uORFs encoding functional proteins should also be classified as polycistronic, but not mRNAs that encode multiple protein isoforms translated from the same ORF.

On 2014 Aug 06, Pavel Baranov commented:

What is a polycistronic mRNA?

This is an interesting research article that provides insights into distinct functions of two proteoforms that differ at their N-termini due to the use of alternative translation initiation starts. Terming the corresponding mRNA bicistronic is, however, somewhat misleading. When we refer to a bacterial mRNA as polycistronic we imply that protein products of distinct coding ORFs are translated from the same transcript. Here the ORF is the same, but the translation initiates at different codons of that ORF. There are several examples of human proteins with truncated or extended N-termini produced from alternative translation initiation starts, see a recently discovered extension of PTEN for a startling example, see Hopkins BD, 2013. True bicistronic mRNAs are rare. Molybdopterin subunits MOCS2A and MOCS2B were reported to be produced from two different but overlapping ORFs at the same mRNA, see Stallmeyer B, 1999. If this were true the corresponding mRNA could be classified as bicistronic, however, further evidence suggested that these proteins are produced from alternatively spliced transcript variants, see Hahnewald R, 2006. A good example of an eukaryotic polycistronic mRNA could be found in Drosophilla tal (aka pri) mRNA that codes for four peptides produced from four distinct ORFs, see Kondo T, 2010. Perhaps, human mRNAs with uORFs encoding functional proteins should also be classified as polycistronic, but not mRNAs that encode multiple protein isoforms translated from the same ORF.