Functional characterization and high-throughput proteomic analysis of interrupted genes in the archaeon Sulfolobus solfataricus.
Sequenced genomes often reveal interrupted coding sequences that complicate the annotation process and the subsequent functional characterization of the genes. In the past, interrupted genes were generally considered to be the result of sequencing errors or pseudogenes, that is, gene remnants with little or no biological importance. However, recent lines of evidence support the hypothesis that these coding sequences can be functional; thus, it is crucial to understand whether interrupted genes are expressed in vivo. We addressed this issue by experimentally demonstrating the existence of functional disrupted genes in archaeal genomes. We discovered previously unknown disrupted genes that have interrupted homologues in distantly related species of archaea. The combination of a RT-PCR strategy with shotgun proteomics demonstrates that interrupted genes in the archaeon Sulfolobus solfataricus are expressed in vivo. In addition, the sequence of the peptides determined by LCMSMS and experiments of in vitro translation allows us to identify a gene expressed by programmed -1 frameshifting. Our findings will enable an accurate reinterpretation of archaeal interrupted genes shedding light on their function and on archaeal genome evolution.
Cobucci-Ponzano
Beatrice
B
Institute of Protein Biochemistry-CNR, Via P. Castellino 111, 80131 Naples, Italy.
Guzzini
Lucia
L
Benelli
Dario
D
Londei
Paola
P
Perrodou
Emmanuel
E
Lecompte
Odile
O
Tran
Diem
D
Sun
Jun
J
Wei
Jing
J
Mathur
Eric J
EJ
Rossi
Mosè
M
Moracci
Marco
M
D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D002853:Chromatography, Liquid; D019766:Genes, Archaeal; D057166:High-Throughput Screening Assays / Q000379:methods*; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D011544:Pseudogenes; D020133:Reverse Transcriptase Polymerase Chain Reaction; D048229:Sulfolobus solfataricus / Q000235:genetics*; D053719:Tandem Mass Spectrometry; D014174:Transketolase / Q000737:chemistry / Q000235:genetics
D019843:Archaeal Proteins; D020543:Proteome; D014174:Transketolase