Large Open Reading Frame Research Articles

Sequence analysis reveals mosaic genome of Aichi virus

Aichi virus is a positive-sense and single-stranded RNA virus, which demonstrated to be related to diarrhea of Children. In the present study, phylogenetic and recombination analysis based on the Aichi virus complete genomes available in GenBank reveal a mosaic genome sequence [GenBank: FJ890523], of which the nt 261-852 region (the nt position was based on the aligned sequence file) shows close relationship with AB010145/Japan with 97.9% sequence identity, while the other genomic regions show close relationship with AY747174/German with 90.1% sequence identity. Our results will provide valuable hints for future research on Aichi virus diversity.Aichi virus is a member of the Kobuvirus genus of the Picornaviridae family [1,2] and belongs to a positive-sense and single-stranded RNA virus. Its presence in fecal specimens of children suffering from diarrhea has been demonstrated in several Asian countries [3-6], in Brazil and German [7], in France [8] and in Tunisia [9]. Some reports showed the high level of seroprevalence in adults [7,10], suggesting the widespread exposure to Aichi virus during childhood.The genome of Aichi virus contains 8,280 nucleotides and a poly(A) tail. The single large open reading frame (nt 713-8014 according to the strain AB010145) encodes a polyprotein of 2,432 amino acids that is cleaved into the typical picornavirus structural proteins VP0, VP3, VP1, and nonstructural proteins 2A, 2B, 2C, 3A, 3B, 3C and 3D [2,11]. Based on the phylogenetic analysis of 519-bp sequences at the 3C-3D (3CD) junction, Aichi viruses can be divided into two genotypes A and B with approximately 90% sequence homology [12]. Although only six complete genomes of Aichi virus were deposited in GenBank at present, mosaic genomes can be found in strains from different countries.

Genomic characterization of a novel dsRNA virus detected in the phytopathogenic fungus Verticillium dahliae Kleb.

Four novel double-stranded RNA segments were detected in a Verticillium dahliae Kleb. strain ( V. dahliae isolate 0-21), a causal fungal agent of Verticillium wilt disease of cotton. Each dsRNA genome segment contains a single large open reading frame (ORF) that encodes a distinctive protein with modest levels of sequence similarities to the corresponding putative proteins in the genus Chrysovirus. These include an RNA-dependent RNA polymerase (RdRp), a coat protein, an undefined replication-related protein and an ovarian tumor domain peptidase. Phylogenetic analysis of the four putative proteins unanimously indicated that they are evolutionarily related to viruses in Chrysovirus. The 5′- and 3′-untranslated regions of the four dsRNAs share highly similar internal sequence and contain conserved sequence stretches of UGAUAAAAAA(/U)UG(/U)AAAAA- (in the 5′-UTR) and -UUUACUACU (in the 3′-UTR), indicating that they have a common virus origin. Indeed, isometric virus-like particles (VLPs) with a diameter of approximately 34 nm were extracted from the fungal mycelia, and the four dsRNA segments were also detected in the virus-like particle (VLP) fraction. These results suggest that the mycovirus with four different dsRNA genome segments from the fungal isolate 0-21 is a new member of the genus Chrysovirus. We named the virus Verticillium dahliae chrysovirus 1 (VdCV1).

Physical and Genomic Characteristics Identify Chicken Proventricular Necrosis Virus (R11/3 Virus) as a Novel Birnavirus

Chicken proventricular necrosis virus (CPNV), isolate R11/3, previously was isolated from transmissible viral proventriculitis-affected chickens and was determined to be the likely etiology of this disease. CPNV was identified as a birnavirus on the basis of virion size and morphology (icosahedral, approximately 75 nm in diameter, nonenveloped); buoyant density in cesium chloride (1.32 g/ml); a genome comprising bisegmented, double-stranded RNA (approximately 3.8 and 3.4 kilobase pairs); and nucleotide sequence analyses. Nucleotide sequencing of CPNV RNA, segment B, identified a single large open reading frame that encodes a 903-amino acid protein. The 903-amino acid protein was identified as the putative VP1, the viral RNA-dependent RNA polymerase (RdRp), on the basis of sequence homologies with other birnavirus VP1 proteins. The CPNV VP1 possessed the unique permuted RdRp sequence motif arrangement characteristic of birnaviruses; however, phylogenetic analyses based on VP1 demonstrated that CPNV is deeply divergent from other birnaviruses.

Complete genome sequence of Celery mosaic virus and its relationship to other members of the genus Potyvirus

The complete genomic sequence of Celery mosaic virus (CeMV) was found to be 9999 nucleotides in length, excluding the 3' poly(A) tail. The genome contains a single large open reading frame encoding a polyprotein of 3181 amino acids. Its genomic organization is typical of potyviruses and contains conserved motifs found in members of the genus Potyvirus. Pairwise comparison of the polyprotein sequences shows that CeMV shares 39.0-71.9% sequence identity with other members of the genus Potyvirus. Phylogenetic analysis based on the polyprotein sequences indicates that CeMV is most closely related to Apium virus Y, and together with Panax virus Y, the three viruses form a distinct clade.

Detection and characterization of two insertion sequences in Vibrio alginolyticus

Vibrio alginolyticus is the dominant Vibrio species and an opportunistic pathogen for marine animals and human. It has been reported to be an important reservoir for virulence genes in aquatic environment. In this study, a total of 187 isolates of V. alginolyticus from fish ponds or coral reefs in the South China Sea were examined for the presence of two insertion sequences, ISVal-1 and ISVal-2. In addition, the characteristics of the two ISs were described. The results showed that 58.82% of the examined isolates contained ISVal-1 and 23.53% had ISVal-2. For both ISVal-1 and ISVal-2, the detection rate for ISs from the fish ponds was lower than that from the coral reef. Sequence analysis revealed that ISVal-1 had the typical features of the IS5 family which contained 19 bp terminal inverted repeats (IRs) and a large open reading frame containing a conserved DDE motif. ISVal-2 possessed genetic structures identical to the IS3 family which had imperfect IRs and two partially overlapping reading frames, OrfA and orfB. A putative helix-turn-helix (HTH) motif, a presumed leucine zipper (LZ) motif, and a conserved DDE motif were contained in the two Orfs. In addition, the data suggested that ISVal-1 and ISVal-2 in V. alginolyticus may be horizontally transferred from other Vibrio species and play certain roles in the virulence of the pathogen. To our knowledge, this is the first report on ISs in V. alginolyticus.

Reverse transcription of the pFOXC mitochondrial retroplasmids of Fusarium oxysporum is protein primed

BackgroundThe pFOXC retroplasmids are small, autonomously replicating DNA molecules found in mitochondria of certain strains of the filamentous fungus Fusarium oxysporum and are among the first linear genetic elements shown to replicate via reverse transcription. The plasmids have a unique clothespin structure that includes a 5'-linked protein and telomere-like terminal repeats, with pFOXC2 and pFOXC3 having iterative copies of a 5 bp sequence. The plasmids contain a single large open reading frame (ORF) encoding an active reverse transcriptase (RT). The pFOXC-RT is associated with the plasmid transcript in a ribonucleoprotein (RNP) complex and can synthesize full-length (-) strand cDNA products. In reactions containing partially purified RT preparations with exogenous RNAs, the pFOXC3-RT has been shown to initiate cDNA synthesis by use of snapped-back RNAs, as well as loosely associated DNA primers.ResultsThe complete sequence of the distantly related pFOXC1 plasmid was determined and found to terminate in 3-5 copies of a 3 bp sequence. Unexpectedly, the majority of (-) strand cDNA molecules produced from endogenous pFOXC1 transcripts were attached to protein. In vitro experiments using partially purified pFOXC3-RT preparations having a single radiolabeled deoxyribonucleotide triphosphate (dNTP) generated a nucleotide-labeled protein that migrated at the size of the pFOXC-RT. The nucleotide preference of deoxynucleotidylation differed between pFOXC3 and pFOXC1 and showed complementarity to the respective 3' terminal repeats. In reactions that include exogenous RNA templates corresponding to the 3' end of pFOXC1, a protein-linked cDNA product was generated following deoxynucleotidylation, suggesting that reverse transcription initiates with a protein primer.ConclusionsThe finding that reverse transcription is protein primed suggests the pFOXC retroplasmids may have an evolutionary relationship with hepadnaviruses, the only other retroelement family known to initiate reverse transcription via a protein primer. Moreover, the similarity to protein-primed linear DNA elements supports models in which the terminal repeats are generated and maintained by a DNA slideback mechanism. The ability of the pFOXC-RT to utilize RNA, DNA and protein primers is unique among polymerases and suggests that the pFOXC plasmids may be evolutionary precursors of a broad range of retroelements, including hepadnaviruses, non-long terminal repeat (non-LTR) retrotransposons and telomerase.

Molecular screening tools to study Arabidopsis transcription factors

In the model plant Arabidopsis thaliana, more than 2000 genes are estimated to encode transcription factors (TFs), which clearly emphasizes the importance of transcriptional control. Although genomic approaches have generated large TF open reading frame (ORF) collections, only a limited number of these genes is functionally characterized, yet. This review evaluates strategies and methods to identify TF functions. In particular, we focus on two recently developed TF screening platforms, which make use of publically available GATEWAY®-compatible ORF collections. (1) The Arabidopsis thaliana TF ORF over-Expression (AtTORF-Ex) library provides pooled collections of transgenic lines over-expressing HA-tagged TF genes, which are suited for screening approaches to define TF functions in stress defense and development. (2) A high-throughput microtiter plate based protoplast trans activation (PTA) system has been established to screen for TFs which are regulating a given promoter:Luciferase construct in planta.

MOLECULAR CHARACTERIZATION AND PHYLOGENETIC STUDIES OF COMPONENT SIX OF BANANA BUNCHY TOP BABUVIRUS (BBTV DNA-6)

Banana and plantations (Musa sp.) are the most fourth most important global food crop after rice, wheat and maize (Piertersen and Thomas, 2001). Banana plantations are subjected to various natural climates but diseases; in particular, viral diseases constitute a major setback to this crop worldwide. Among viral infections, Banana Bunchy Top Virus (BBTV) is the most serious virus affecting banana in Egypt as well as the world (Dale, 1987; Othman et al., 1996; Harish et al., 2008) and is responsible for the massive reduction in banana crop yield. The symptoms of BBTD are yellowing, dark green streaks on midrib, petioles, extending down into pseudostem, dark green dots and dashes along the minor leaf veins, stunted growth with bunchy top appearance (Magee, 1940; Vetten et al., 2005). BBTV belong to genus Babuvirus and family Nanoviridae with isometric particles of 18-20 nm in diameter. It is naturally transmitted, in persistence manner, by banana aphid Pentalonia nigronervosa (Hu et al., 1996). BBTV is identified as a phloem-limited virus used to occur in low concentration in banana-infected plants (Harding et al., 1991 and 2000). The virus has a single coat protein with molecular weight (Mr.) of about 20-21kDa (Dietzgen and Thomas, 1991; Harding et al., 1991; Sadik et al., 1999).The viron consists of at least six components of circular ssDNA each component of the BBTV genome contains one large (monocistronic) transcriptional active open reading frame (ORF) except BBTV DNA 1 contains two ORFs one master replication protein (Rep) ORF (Hafner et al., 1997) and a small ORF internal to major ORF (Beetham et al., 1997). Additionally, two conserved region: stem-loop common region (CR-SL) and major common region (CR-M), potential TATA box at 3 ' of the stem-loop and polyadenylation signal (Burns et al., 1995; Beetham et al., 1997 and 1999).The nucleotide sequence of the component six of Australian isolate BBTV has 1089 nt. (Burns et al., 1995). The encoded protein of BBTV DNA-6 is a nuclear shuttle protein and is preferentially targeted to the nucleus when expressed alone however in the presence of the movement protein it targeted to the cell periphery (Wanitchakorn et al., 2000a).The aim of this study is to amplify, clone and sequence the full length of BBTV DNA-6 and compare it with other published sequences of the corresponding sequence.

Molecular and Biological Characterization of Chinese Sacbrood Virus LN Isolate

Chinese sacbrood virus (CSBV) was purified from diseased insects, and its genome was cloned and sequenced. The genomic RNA of CSBV is 8863 nucleotides in length and contains a single large open reading frame encoding a 319.614 kDa polyprotein. The coding sequence is flanked by a 178-nucleotide 5′ nontranslated leader sequence and a 142-nucleotide 3′ nontranslated region, followed a poly(A) tail. Four major structural proteins, VP1,VP2, VP3 and VP4, were predicted in the N-teminal of the polyprotein. The C-terminal part of the polyprotein contains sequence motifs which is a typical and well-characterized picornavirus nonstructural proteins: an RNA helicase, a chymotrypsin-like 3C protease, and an RNA-dependent RNA polymerase. Genetic analysis shows that the CSBV-LN had a 13-amino-acid deletion at amino acid positions 710–719 and 727–729 in comparison with CSBV-GZ and SBV-UK, and the SBV-UK had a 7-amino-acid deletion at amino acid positions 2124–2132 in comparison with CSBV-GZ and CSBV-LN, and the CSBV-GZ and CSBV-LN had a 6-amino-acid deletion at amino acid positions 2143–2150 in comparison with SBV-UK. Phylogenetic analysis using RdRp of selected picorna-like viruses shows that CSBV/SBV and Deformed Wing Virus (DWV) tend to group together, which possesses an RNA of similar size and gene order.

Mass Spectrometric Analysis of Synapsins in Drosophila melanogaster and Identification of Novel Phosphorylation Sites

Synapsins are synaptic vesicle-associated phosphoproteins that play a major role in the fine regulation of neurotransmitter release. In Drosophila, synapsins are required for complex behavior including learning and memory. Synapsin isoforms were immunoprecipitated from homogenates of wild-type Drosophila heads using monoclonal antibody 3C11. Synapsin null mutants (Syn(97)) served as negative controls. The eluted proteins were separated by SDS-PAGE and visualized by silver staining. Gel pieces picked from five bands specific for wild type were analyzed by nano-LC-ESI-MS/MS following multienzyme digestion (trypsin, chymotrypsin, AspN, subtilisin, pepsin, and proteinase K). The protein was unambiguously identified with high sequence coverage (90.83%). A number of sequence conflicts were observed and the N-terminal amino acid was identified as methionine rather than leucine expected from the cDNA sequence. Several peptides from the larger isoform demonstrated that the in-frame UAG stop codon at position 582 which separates two large open reading frames is read through by tRNAs for lysine. Seven novel phosphorylation sites in Drosophila synapsin were identified at Thr-86, Ser-87, Ser-464, Thr-466, Ser-538, Ser-961, and Tyr-982 and verified by phosphatase treatment. No phosphorylation was observed at the conserved PKA/CaM kinase-I/IV site (RRFS, edited to RGFS) in domain A or a potential PKA site near domain E.

The complete genome sequence of a Passion fruit woodiness virus isolate from Australia determined using deep sequencing, and its relationship to other potyviruses

The complete genome sequence (9,858 nucleotides) of the Passion fruit woodiness virus isolate MU-2 was determined using Illumina sequencing. The large open reading frame (ORF) encodes a polyprotein containing 3,086 amino acids, with an AUG start codon and UAA stop codon. The polyprotein yielded 11 proteins (P1, HC-Pro, P3, PIPO, 6K1, CI, 6K2, NIa-VPg, NIa-Pro, NIb and CP). Putative cleavage sites between them were identified by sequence comparison to those of other known potyviruses. Accuracy of the genome sequence information was provided by 42-1691-fold sequence coverage, and viral RNA accounted for 7.38% of total polyadenylated RNA from the host plant.

Clostridium perfringens bacteriophages ΦCP39O and ΦCP26F: genomic organization and proteomic analysis of the virions.

Poultry intestinal material, sewage and poultry processing drainage water were screened for virulent Clostridium perfringens bacteriophages. Viruses isolated from broiler chicken offal washes (O) and poultry feces (F), designated ΦCP39O and ΦCP26F, respectively, produced clear plaques on host strains. Both bacteriophages had isometric heads of 57 nm in diameter with 100-nm non-contractile tails characteristic of members of the family Siphoviridae in the order Caudovirales. The double-strand DNA genome of bacteriophage ΦCP39O was 38,753 base pairs (bp), while the ΦCP26F genome was 39,188 bp, with an average GC content of 30.3%. Both viral genomes contained 62 potential open reading frames (ORFs) predicted to be encoded on one strand. Among the ORFs, 29 predicted proteins had no known similarity while others encoded putative bacteriophage capsid components such as a pre-neck/appendage, tail, tape measure and portal proteins. Other genes encoded a predicted DNA primase, single-strand DNA-binding protein, terminase, thymidylate synthase and a transcription factor. Potential lytic enzymes such as a fibronectin-binding autolysin, an amidase/hydrolase and a holin were encoded in the viral genomes. Several ORFs encoded proteins that gave BLASTP matches with proteins from Clostridium spp. and other Gram-positive bacterial and bacteriophage genomes as well as unknown putative Collinsella aerofaciens proteins. Proteomics analysis of the purified viruses resulted in the identification of the putative pre-neck/appendage protein and a minor structural protein encoded by large open reading frames. Variants of the portal protein were identified, and several mycobacteriophage gp6-like protein variants were detected in large amounts relative to other virion proteins. The predicted amino acid sequences of the pre-neck/appendage proteins had major differences in the central portion of the protein between the two phage gene products. Based on phylogenetic analysis of the large terminase protein, these phages are predicted to be pac-type, using a head-full DNA packaging strategy.

Biological characterization and complete genomic sequence of Apium virus Y infecting celery

A celery isolate of Apium virus Y (ApVY-Ce) from diseased plants in a commercial field in California was characterized. The experimental host range of the virus included 13 plant species in the families Apiaceae, Chenopodiaceae and Solanaceae. Almost all infected plant species showed foliar chlorosis and distortion or severe stunting and systemic chlorosis. ApVY-Ce was transmitted to all 10 host species in the Apiaceae by green peach aphids. It reacted with the potyvirus group antibody and Celery mosaic virus (CeMV) antiserum. The complete genomic sequence of ApVY-Ce was determined to be 9917 nucleotides, excluding the 3′ poly(A) tail, and it comprises a large open reading frame encoding a polyprotein of 3184 amino acid residues. Its genomic organization is typical of potyviruses, and contains conserved motifs found in the genus Potyvirus. Comparisons with available genomic sequences of other potyviruses indicate that ApVY-Ce shares 26.1–52.9% identities with species of the existing genera and unassigned viruses in the Potyviridae at the polyprotein sequence level. Extensive phylogenetic analysis based on the 3′-partial sequences confirms that ApVY-Ce is most closely related to CeMV and is a distinct species of the genus Potyvirus.

AN UNPRECEDENTED GROUP-II INTRON FROM CHLAMYDOMONAS SUBCAUDATA (CHLOROPHYCEAE) THAT HAS TWO LARGE OPEN READING FRAMES-POTENTIALLY ENCODING A REVERSE TRANSCRIPTASE-MATURASE-ENDONUCLEASE AND A NOVEL PROTEIN1

Group-II introns are selfish ribozymes that may have given rise to nuclear mRNA introns. Approximately 1,000 of these introns—derived from organelles, bacteria, and archaea—have been defined as either having no open reading frame (ORF) or encoding a single large protein, which is nearly always a variant of a reverse transcriptase-maturase-endonuclease (RT-Mat-En). While investigating intron ribozymes in cold-tolerant Chlamydomonas spp., we discovered an unusually large (3.9 kb) group-II intron in the psbA gene of Chlamydomonas subcaudata N. Wille, Csu.psbA. Reverse transcriptase-PCR (RT-PCR) analysis showed that Csu.psbA is efficiently spliced in vivo and confirmed the predicted splice sites. The extreme size of Csu.psbA is due to two large ORFs in domain IV of the predicted secondary structure. ORF1 encodes a typical RT-Mat-En protein (70 kDa), although it has an unusual start codon (ACG). ORF2, however, encodes a potentially novel protein (44 kDa) that is predicted to have a transmembrane domain, immediately following an N-terminal thylakoid-targeting peptide, and to bind nucleic acids. BLAST analyses suggest that both ORFs are of bacterial origin and that ORF2 may have a TRKA domain. Csu.psbA is the first group-II intron reported to have two large, distinct ORFs and raises the possibility of identifying novel intron-encoded functions.

Time-dependent increase in ribosome processivity

We created a novel tripartite reporter RNA to separately and simultaneously examine ribosome translation rates at the 5′- and 3′-ends of a large open reading frame (ORF) in vitro in HeLa cell lysates. The construct contained Renilla luciferase (RLuc), β-galactosidase and firefly luciferase (FLuc) ORFs linked in frame and separated by a viral peptide sequence that causes cotranslational scission of emerging peptide chains. The length of the ORF contributed to low ribosome processivity, a low number of initiating ribosomes completing translation of the entire ORF. We observed a time-dependent increase in FLuc production rate that was dependent on a poly(A) tail and poly(A)-binding protein, but was independent of eIF4F function. Stimulation of FLuc production occurred earlier on shorter RNA templates. Cleavage of eIF4G at times after ribosome loading on templates occurred did not cause immediate cessation of 5′-RLuc translation; rather, a delay was observed that shortened when shorter templates were translated. Electron microscopic analysis of polysome structures in translation lysates revealed a time-dependent increase in ribosome packing and contact that correlated with increased processivity on the FLuc ORF. The results suggest that ORF transit combined with PABP function contribute to interactions between ribosomes that increase or sustain processivity on long ORFs.

Complete genomic sequence of a border disease virus isolated from Pyrenean chamois

This report describes the full-length genome sequence of the pestivirus strain H2121 which was recently isolated from Pyrenean chamois and typed as Border disease virus (BDV) genotype 4. Comparison with full-length genomic sequences of the approved pestivirus species Bovine viral diarrhea virus-1 (BVDV-1), BVDV-2, BDV, and Classical swine fever virus, the tentative species represented by strain Giraffe-1, as well as the atypical pestivirus strain Th/04_KhonKaen confirmed that the chamois pestivirus strain is most similar to BDV. The viral genome of H2121 is 12,305 nucleotides long and contains one large open reading frame. The latter encodes a polyprotein consisting of 3899 amino acids and is flanked with 376 nucleotides long 5′ untranslated region (UTR) and 229 nt long 3′ UTR. The genome organization of the chamois virus is reminiscent to that of other pestiviruses. Compared to other BDV strains including BDV-1 strain X818 and BDV-2 strain Reindeer-1, the 5′ UTR and ORF of the chamois virus are very similar in length, while the 3′ UTR of H2121 is 31–44 nucleotides shorter. In contrast to other BDV strains, the genome of the chamois virus contains a unique four amino acid insertion at the N-terminus of NS2.

Redefining the Genetics of Murine Gammaherpesvirus 68 via Transcriptome-Based Annotation

Viral genetic studies typically focus on large open reading frames (ORFs) identified during genome annotation (ORF-based annotation). Here we describe tools for examining viral gene expression nucleotide by nucleotide across the genome. Using these tools on the 119,450 base pair (bp) genome of murine gammaherpesvirus 68 (gammaHV68) allowed us to establish that gammaHV68 RNA expression was significantly more complex than predicted from ORF-based annotation, including over 73,000 nucleotides of unexpected transcription within 30 expressed genomic regions (EGRs). Approximately 90% of this RNA expression was antisense to genomic regions containing known large ORFs. We verified the existence of previously undefined transcripts in three EGRs and determined which parts of the transcriptome depend on protein or viral DNA synthesis. This study redefines the genetic map of gammaHV68, indicating that herpesviruses contain significantly more genetic complexity than predicted from ORF-based genome annotations, and provides alternative tools and approaches for viral genetic studies.

Genetic heterogeneity found in the replicase gene of poinsettia mosaic virus isolates

The complete nucleotide sequences of five isolates of poinsettia mosaic virus (PnMV) from Japan (JN, JO1, JO2, JO4, and JO5) were determined. These isolates contained a single large open reading frame in their genomes and shared 96.6-97.8% identity at the nucleotide level and 91.3-98.1% identity at the amino acid level with two previously reported European isolates. Interestingly, the JO isolates were found to possess eight common translational frameshift sites in the interdomain region between the methyltransferase and protease domains, resulting in considerable variation in the interdomain region compared to the other isolates. This suggests that PnMV might have evolved by creating variations in its genome by such translational frameshifts.

Insoluble DISC1–Boymaw fusion proteins generated by DISC1 translocation

Insoluble DISC1–Boymaw fusion proteins generated by DISC1 translocation

Complete nucleotide sequence and taxonomy of Sugarcane streak mosaic virus, member of a novel genus in the family Potyviridae

The complete genomic sequence of a Pakistani isolate of Sugarcane streak mosaic virus (SCSMV-PAK) is determined to be 9782 nucleotides in length, excluding the 3' poly(A) tail, and it comprises a large open reading frame encoding a polyprotein of 3130 amino acid residues. The deduced polyprotein is likely to be cleaved at nine putative protease sites by three viral proteases to ten mature proteins. Conserved motifs of orthologous proteins of other potyviruses are identified in corresponding positions of SCSMV-PAK. The genomic organization is virtually identical to the genera Ipomovirus, Potyvirus, Rymovirus, and Tritimovirus in the family Potyviridae. Sequence analyses indicate that the SCSMV-PAK genomic sequence is different from those of Sugarcane mosaic virus and Sorghum mosaic virus, two viruses with very similar symptoms and host range to SCSMV-PAK. SCSMV-PAK shares 52.7% identity with Triticum mosaic virus (TriMV) and 26.4-31.5% identities with species of the existing genera and unassigned viruses in the Potyviridae at the polyprotein sequence level. Phylogenetic analyses of the polyprotein and deduced mature protein amino acid sequences reveal that SCSMV, together with TriMV, forms a distinct group in the family at the genus level. Therefore, SCSMV should represent a new genus, Susmovirus, in the Potyviridae.