Characterization of the ovine ribosomal protein SA gene and its pseudogenes

Abstract

Background: The ribosomal protein SA (RPSA), previously named 37-kDa laminin receptor precursor/67-kDa laminin receptor (LRP/LR) is a multifunctional protein that plays a role in a number of pathological processes, such as cancer and prion diseases. In all investigated species, RPSA is a member of a multicopy gene family consisting of one full length functional gene and several pseudogenes. Therefore, for studies on RPSA related pathways/pathologies, it is important to characterize the whole family and to address the possible function of the other RPSA family members. The present work aims at deciphering the RPSA family in sheep.

Results: In addition to the full length functional ovine RPSA gene, 11 other members of this multicopy gene family, all processed pseudogenes, were identified. Comparison between the RPSA transcript and these pseudogenes shows a large variety in sequence identities ranging from 99% to 74%. Only one of the 11 pseudogenes, i.e. RPSAP7, shares the same open reading frame (ORF) of 295 amino acids with the RPSA gene, differing in only one amino acid. All members of the RPSA family were annotated by comparative mapping and fluorescence in situ hybridization (FISH) localization. Transcription was investigated in the cerebrum, cerebellum, spleen, muscle, lymph node, duodenum and blood, and transcripts were detected for 6 of the 11 pseudogenes in some of these tissues.

Conclusions: In the present work we have characterized the ovine RPSA family. Our results have revealed the existence of 11 ovine RPSA pseudogenes and provide new data on their structure and sequence. Such information will facilitate molecular studies of the functional RPSA gene taking into account the existence of these pseudogenes in the design of experiments. It remains to be investigated if the transcribed members are functional as regulatory non-coding RNA or as functional proteins.

Figure 1

Comparative mapping of the region of RPSA in sheep and cattle. The ovine BAC mini-contig is drawn in part A. Triangles represent BAC end sequences; pointing towards the 3'-end of the BAC clone. Black triangles represent BES from which primers were designed to construct the mini-contig. White triangles are BES from which it was impossible to design STS-primers. Black squares show overlaps between BES and other BAC clones. Black circles represent genes annotated by PCR. Annotated sequences are shown in a plane map in part B. The position and orientation of the genes present in the syntenic region of Bos taurus are represented with arrows (C).

Figure 2

Comparative mapping of the region of 11 RPSA pseudogenes in sheep and cattle. The ovine BAC mini-contigs are drawn in part A. Triangles represent BAC end sequences; pointing towards the 3'-end of the BAC clone. Black triangles represent BES from which primers were designed to construct the mini-contigs. White triangles are BES from which it was impossible to design STS-primers. Encircled triangles represent BES that are not annotated. Black squares show overlaps between BES and other BAC clones. Black circles represent genes annotated by PCR. Annotated sequences are shown in a plane map in part B. The position and orientation of the genes present in the syntenic region of Bos taurus are represented with arrows (C).

Figure 3

Schematic overview of the RPSA (pseudo)genes. The genomic structure of the ovine RPSA gene is drawn in part A, the mRNA of the RPSA gene is drawn in part B. Part C represents the genomic structure of the different RPSA pseudogenes. The squares represent exons and the lines stand for introns. The coding sequence (CDS) is drawn in yellow; the untranslated sequences in green. The blue squares are parts of the pseudogene sequence that are analogous with the exons of the RPSA mRNA. The pink squares symbolize interspersed sequences and the white gaps deletions. SNORA62 is represented as a black square. Start codons and stop codons, analogous with the ones of RPSA, are represented by a dotted line.

Figure 4

Alignment of the snoRNAs in RPSA, RPSAP8 and RPSAP9. The ACA-box, H-box and 28S rRNA U3830 and U3832 PU guide are highlighted in yellow.

Figure 5

Transcription profile of the RPSA gene family members. Marker (M) is the Hyperladder V or IV (Bioline). Samples are cerebrum (Cbu), cerebellum (Cbe), spleen (Sp), muscle (Mu), lymph node (Ln), duodenum (Dd), blood (Bl) genomic or BAC DNA (+) and water (-).