αr15 is a family of bacterial small non-coding RNAs with representatives in a broad group of α-proteobacteria from the order Rhizobiales. The first members of this family (smr15C1 and smrC15C2) were found tandemly arranged in the same intergenic region (IGR) of the Sinorhizobium meliloti 1021 chromosome (C). Further homology and structure conservation analysis have identified full-length Smr15C1 and Smr15C2 homologs in several nitrogen-fixing symbiotic rhizobia (i.e. R. leguminosarum bv. viciae, R. leguminosarum bv. trifolii, R. etli, and several Mesorhizobium species), in the plant pathogens belonging to Agrobacterium species (i.e. A. tumefaciens, A. vitis, A. radiobacter, and Agrobacterium H13) as well as in a broad spectrum of Brucella species (B. ovis, B. canis, B. abortus and B. microtis, and several biovars of B. melitensis). The Smr15C1 (115 nt) and Smr15C2 (121 nt) homologs are also encoded in tandem within the same IGR region of Rhizobium and Agrobacterium species, whereas in Brucella species the αr15C loci are spread in the IGRs of Chromosome I. Moreover, this analysis also identified a third αr15 loci in extrachromosomal replicons of the mentioned nitrogen-fixing α-proteobacteria and in the Chromosome II of Brucella species. αr15 RNA species are 99-121 nt long (Table 1) and share a well defined common secondary structure consisting of three stem loops (Figure 1). The transcripts of the αr15 family can be catalogued as trans-acting sRNAs encoded by independent transcription units with recognizable promoter and transcription termination signatures within intergenic regions (IGRs) of the α-proteobacterial genomes (Figure 5).
Discovery and structure Smr15C1 y Smr15C2 sRNAs were described by del Val et al., as a result of a computational comparative genomic approach in the intergenic regions (IGRs) of the reference S. meliloti 1021 strain (http://iant.toulouse.inra.fr/bacteria/annotation/cgi/rhime.cgi). Although the primary nucleotide sequence of Smr15C1 y Smr15C2 showed high similarity (84% identity), specific probes for each sRNA could be designed which detected transcripts of different size and expression profiles.
TAP-based 5'-RACE experiments mapped the Smr15C1 and Smr15C2 transcription start sites (TSS) in the S. meliloti 1021 genome (http://iant.toulouse.inra.fr/bacteria/annotation/cgi/rhime.cgi). The Smr15C1 TSS was mapped to the chromosomal position 1698731 nt and the TSS of Smr15C2 to the nt 1698937. The 3'-ends were assumed to be located at the 1698617 nt and 1698817 nt respectively, matching the last residue of the consecutive Us stretch of a bona fide Rho-independent terminator (Figure 5). Parallel and later studies, in which Smr15C1 and Smr15C2 transcripts are referred to as two copies of sra41 or Sm3/Sm3', independently confirmed the expression of these sRNAs in S. melilloti and in its closely related strain 2011. Recent deep sequencing-based characterization of the small RNA fraction (50-350 nt) of S. meliloti 2011 also revealed the expression of Smr15C1 and Smr15C2, here referred to as SmelC411 and SmelC412 respectively, mapping the 5'- and 3'-ends of the full-length transcripts to essentially the same positions as del Val et al. in the S. meliloti 1021 chromosome. However, this study identified an additional TSS for Smr15C2 at position 1698948.
The nucleotide sequences of Smr15C1 and Smr15C2 were initially used as query to search against the Rfam database (version 10.0; http://rfam.xfam.org). This search revealed partial homology of both transcripts, restricted to the second hairpin and the Rho-independent terminator, to the RF00519 family of RNAs known as suhB (http://rfam.xfam.org/family/RF00519). However, no structural homologs of the full-length sRNAs were found in this database.
Both S.melilloti αr15 sRNAs were also BLASTed with default parameters against all the currently available bacterial genomes (1,615 sequences at 20 April 2011; https://www.ncbi.nlm.nih.gov). The regions exhibiting significant homology to the query sequence (78-89% similarity) were extracted to create a Covariance Model (CM) from a seed alignment using Infernal (version1.0) (Figure 2). This CM was used in a further search for new members of the αr15 family in the existing bacterial genomic databases.
+ Table 1: Smr15C1 and Smr15C2 homologs in other symbionts and pathogens CM model Name GI accession number begin end strand %GC length Organism αr15 Smr15C1 gi15963753refNC_003047.1 1698617 1698731 - 54 115 Sinorhizobium meliloti 1021 αr15 Smr15C2 gi15963753refNC_003047.1 1698817 1698937 - 50 121 Sinorhizobium meliloti 1021 αr15 Smr15A gi16262453refNC_003037.1 552873 552984 + 51 112 Sinorhizobium meliloti 1021 plasmid pSymA αr15 Smedr15C1 gi150395228refNC_009636.1 1337011 1337126 - 53 116 Sinorhizobium medicae WSM419 chromosome αr15 Smedr15C2 gi150395228refNC_009636.1 1337212 1337331 - 50 120 Sinorhizobium medicae WSM419 chromosome αr15 Smedr15p03 gi150378263refNC_009622.1 40054 40165 - 52 112 Sinorhizobium medicae WSM419 plasmid pSMED03 αr15 Sfr15C1 gi227820587refNC_012587.1 1612511 1612626 - 59 116 Sinorhizobium fredii NGR234 chromosome αr15 Sfr15C2 gi227820587refNC_012587.1 1612711 1612830 - 51 120 Sinorhizobium fredii NGR234 chromosome αr15 Sfr15b gi227818258refNC_012586.1 134078 134190 - 55 113 Sinorhizobium fredii NGR234 plasmid pNGR234b αr15 Atr15C1 gi159184118refNC_003062.2 2163254 2163370 + 53 117 Agrobacterium tumefaciens str. C58 chromosome circular αr15 Atr15C2 gi159184118refNC_003062.2 2163454 2163554 + 57 101 Agrobacterium tumefaciens str. C58 chromosome circular αr15 AH13r15C1 gi325291453refNC_015183.1 2112823 2112939 + 52 117 Agrobacterium sp. H13-3 chromosome αr15 AH13r15C2 gi325291453refNC_015183.1 2113023 2113121 + 54 99 Agrobacterium sp. H13-3 chromosome αr15 AH13r15a gi325168279refNC_015184.1 211698 211807 - 53 110 Agrobacterium sp. H13-3 plasmid pAspH13-3a αr15 ReCIATr15C1 gi190889639refNC_010994.1 3155217 3155332 + 51 116 Rhizobium etli CIAT 652 αr15 ReCIATr15C2 gi190889639refNC_010994.1 3155440 3155555 + 48 116 Rhizobium etli CIAT 652 αr15 ReCIATr15pC gi190894340refNC_010997.1 941345 941452 + 53 108 Rhizobium etli CIAT 652 plasmid pC αr15 ReCIATr15B gi190893983refNC_010996.1 187927 188041 - 50 115 Rhizobium etli CIAT 652 plasmid pB αr15 Arr15CI1 gi222084201refNC_011985.1 2506215 2506331 + 52 117 Agrobacterium radiobacter K84 chromosome 1 αr15 Arr15CI2 gi222084201refNC_011985.1 2506418 2506534 + 54 117 Agrobacterium radiobacter K84 chromosome 1 αr15 Arr15CII gi222080781refNC_011983.1 1011511 1011624 - 57 114 Agrobacterium radiobacter K84 chromosome 2 αr15 Rlt2304r15C1 gi209547612refNC_011369.1 2770612 2770727 + 50 116 Rhizobium leguminosarum bv. trifolii WSM2304 chromosome αr15 Rlt2304r15C2 gi209547612refNC_011369.1 2770835 2770949 + 50 115 Rhizobium leguminosarum bv. trifolii WSM2304 chromosome αr15 Avr15CI1 gi222147015refNC_011989.1 2608532 2608647 + 54 116 Agrobacterium vitis S4 chromosome 1 αr15 Avr15CI2 gi222147015refNC_011989.1 2608739 2608839 + 46 101 Agrobacterium vitis S4 chromosome 1 αr15 Avr15Atc gi222083145refNC_011984.1 122624 122736 - 50 113 Agrobacterium vitis S4 plasmid pAtS4c αr15 Avr15Ate gi222102412refNC_011981.1 198928 199039 + 51 112 Agrobacterium vitis S4 plasmid pAtS4e αr15 Avr15Ti gi222080117refNC_011982.1 52286 52397 - 57 112 Agrobacterium vitis S4 plasmid pTiS4 αr15 Rlvr15C1 gi116249766refNC_008380.1 3605490 3605605 + 51 116 Rhizobium leguminosarum bv. viciae 3841 αr15 Rlvr15C2 gi116249766refNC_008380.1 3605714 3605829 + 48 116 Rhizobium leguminosarum bv. viciae 3841 αr15 Rlvr15p10 gi116254467refNC_008381.1 138799 138912 + 56 114 Rhizobium leguminosarum bv. viciae 3841 plasmid pRL10 αr15 Rlvr15p11 gi116255200refNC_008384.1 567053 567166 - 53 114 Rhizobium leguminosarum bv. viciae 3841 plasmid pRL11 αr15 Rlt1325r15C1 gi241202755refNC_012850.1 2981275 2981390 + 50 116 Rhizobium leguminosarum bv. trifolii WSM1325 αr15 Rlt1325r15C2 gi241202755refNC_012850.1 2981499 2981613 + 50 115 Rhizobium leguminosarum bv. trifolii WSM1325 αr15 Rlt1325r15p02 gi241666492refNC_012858.1 36176 36289 + 51 114 Rhizobium leguminosarum bv. trifolii WSM1325 plasmid pR132502 αr15 ReCFNr15C1 gi86355669refNC_007761.1 3117667 3117782 + 50 116 Rhizobium etli CFN 42 αr15 ReCFNr15C2 gi86355669refNC_007761.1 3117890 3118004 + 50 115 Rhizobium etli CFN 42 αr15 ReCFNr15d gi89255298refNC_004041.2 172760 172874 - 50 115 Rhizobium etli CFN 42 symbiotic plasmid p42d αr15 ReCFNr15a gi86359705refNC_007762.1 157296 157409 + 56 114 Rhizobium etli CFN 42 plasmid p42a αr15 Mlr15a gi13488050refNC_002679.1 65044 65154 - 51 111 Mesorhizobium loti MAFF303099 plasmid pMLa αr15 Bcr15CII gi161620094refNC_010104.1 707465 707572 + 56 108 Brucella canis ATCC 23365 chromosome II αr15 Bcr15CI1 gi161617991refNC_010103.1 1379297 1379398 - 51 102 Brucella canis ATCC 23365 chromosome I αr15 Bcr15CI2 gi161617991refNC_010103.1 1451969 1452087 - 50 119 Brucella canis ATCC 23365 chromosome I αr15 Bs23445r15CI1 gi163842277refNC_010169.1 1401085 1401186 - 51 102 Brucella suis ATCC 23445 chromosome I αr15 Bs23445r15CI2 gi163842277refNC_010169.1 1473791 1473909 - 50 119 Brucella suis ATCC 23445 chromosome I αr15 Bs23445r15CII gi163844199refNC_010167.1 696081 696188 + 56 108 Brucella suis ATCC 23445 chromosome II αr15 Bm16Mr15CI gi17986284refNC_003317.1 607684 607785 + 51 102 Brucella melitensis bv. 1 str. 16M chromosome I αr15 Bm16Mr15CII gi17988344refNC_003318.1 589501 589608 - 56 108 Brucella melitensis bv. 1 str. 16M chromosome II αr15 BaS19r15CII gi189022234refNC_010740.1 508055 508162 - 56 108 Brucella abortus S19 chromosome 2 αr15 BaS19r15CI1 gi189023268refNC_010742.1 1396794 1396895 - 51 102 Brucella abortus S19 chromosome 1 αr15 BaS19r15CI2 gi189023268refNC_010742.1 1469407 1469525 - 50 119 Brucella abortus S19 chromosome 1 αr15 Bm23457r15CII gi225685871refNC_012442.1 687290 687397 + 56 108 Brucella melitensis ATCC 23457 chromosome II αr15 Bm23457r15CI gi225851546refNC_012441.1 1400641 1400742 - 51 102 Brucella melitensis ATCC 23457 chromosome I αr15 Bs1330r15CII gi56968493refNC_004311.2 708185 708292 + 56 108 Brucella suis 1330 chromosome II αr15 Bs1330r15CI1 gi56968325refNC_004310.3 1380381 1380482 - 51 102 Brucella suis 1330 chromosome I αr15 Bs1330r15CI2 gi56968325refNC_004310.3 1453011 1453129 - 50 119 Brucella suis 1330 chromosome I αr15 Ba19941r15CI1 gi62288991refNC_006932.1 1398464 1398565 - 51 102 Brucella abortus bv. 1 str. 9-941 chromosome I αr15 Ba19941r15CI2 gi62288991refNC_006932.1 1471073 1471191 - 50 119 Brucella abortus bv. 1 str. 9-941 chromosome I αr15 Ba19941r15CII gi62316961refNC_006933.1 508851 508958 - 56 108 Brucella abortus bv. 1 str. 9-941 chromosome II αr15 Bmar15CII gi83268957refNC_007624.1 508839 508946 - 56 108 Brucella melitensis biovar Abortus 2308 chromosome II αr15 Bmar15CI1 gi82698932refNC_007618.1 1395614 1395715 - 51 102 Brucella melitensis biovar Abortus 2308 chromosome I αr15 Bmar15CI2 gi82698932refNC_007618.1 1468227 1468345 - 50 119 Brucella melitensis biovar Abortus 2308 chromosome I αr15 Bor15CI1 gi148558820refNC_009505.1 1387928 1388029 - 50 102 Brucella ovis ATCC 25840 chromosome I αr15 Bor15CI2 gi148558820refNC_009505.1 1460506 1460624 - 50 119 Brucella ovis ATCC 25840 chromosome I αr15 Bor15CII gi148557829refNC_009504.1 709415 709524 + 54 110 Brucella ovis ATCC 25840 chromosome II αr15 Bmir15CII gi256014795refNC_013118.1 709102 709209 + 56 108 Brucella microti CCM 4915 chromosome 2 αr15 Bmir15CI1 gi256368465refNC_013119.1 1387776 1387877 - 51 102 Brucella microti CCM 4915 chromosome 1 αr15 Bmir15CI2 gi256368465refNC_013119.1 1461298 1461416 - 50 119 Brucella microti CCM 4915 chromosome 1 αr15 Oar15CI gi153007346refNC_009667.1 1751482 1751598 + 49 117 Brucella anthropi ATCC 49188 chromosome 1 αr15 Oar15CII gi153010078refNC_009668.1 1270083 1270191 + 56 109 Brucella anthropi ATCC 49188 chromosome 2 αr15 Oar15p02 gi153011934refNC_009670.1 22208 22320 + 54 113 Brucella anthropi ATCC 49188 plasmid pOANT02