a blast search restricted to mammalian protein sequences was performed through the ncbi web page ( http://www.ncbi.nlm.nih.gov/blast ) by using the nipah virus matrix protein sequence as the query . one sequence , accession number aak76747 , with homology to the nipah virus matrix protein , was identified . this protein , called angrem52 ( for angiotensin ii - induced , renal mesangial cell gene 52 ) , displays 53% amino acid identity and 73% amino acid similarity over 337 amino acids ( aa ) to the nipah virus matrix protein ( data not shown ) . the angrem52 protein sequence is derived from a theoretical translation of an open reading frame ( orf ) from nucleotides ( nt ) 16 to 1038 within the 3170-nt long angrem52 nucleotide sequence ( genbank accession no . this notable homology suggests that angrem52 is actually a paramyxovirus m gene ( figure 1a ) . angrem52 and a ) gene positions of a generic paramyxovirus and predicted genome position of angrem104 ( top ) , the phosphoprotein ( p ) gene , angrem52 ( bottom ) , the matrix protein ( m ) and fusion protein ( f ) genes . a potential editing site ( nucleotides 783795 ) , which might allow production from the opmv p gene of v and w / d proteins , is shown in genomic ( negative ) sense aligned with the proposed editing sites of nipah virus ( nc_002728 ) ( 1 ) and hendra viruses ( nc_001906 ) . the full - length p open reading frame ( orf ) was obtained by inserting an additional nucleotide in the reported angrem104 sequence ( see text ) . angrem52 is predicted to be a  read - through  product of the m and f genes , a novel paramyxovirus . the full - length f orf was obtained by making 5 changes to the reported angrem52 sequence ( see text ) . putative gene - end , intergenic ( ig ) , and gene - start transcription regulatory signals lying between opmv m and f genes are shown aligned to the corresponding signals from nipah and hendra virus ( shown in genomic sense ) . the sequences surrounding the f protein cleavage sites , including most fusion peptides , of several paramyxoviruses , including putative opmv , were aligned by using the alignx program of the vector nti6 software package . c ) putative genome organization of the putative opmv p gene , allowing translation of p , v , w , and c orfs . d ) alignment of cysteine - rich carboxy - termini of the putative opmv and nipah virus v proteins . the conserved carboxy - terminal regions of the v proteins were aligned by using the alignx program of the vector nti6 software package . conserved residues are indicated in red , except for conserved cysteines , which are in blue . underlined residues are conserved among all paramyxovirus v proteins . upon further analysis of sequences within the angrem52 cdna downstream of the putative matrix gene , several relatively short orfs were found to encode peptides with homology to paramyxovirus fusion ( f ) proteins . modification of the reported angrem52 sequence in several positions yields what appears to be a full - length or near full - length paramymyxovirus fusion ( f ) protein gene , which would be separated from the m orf by 355 nt ( figure 1a ) . specifically , the f orf within the original angrem52 sequence begins at position 1393 but appears to terminate prematurely with a stop codon at 2118 . to obtain what appears to be a f orf , several modifications were made to the reported sequence in order to to incorporate the additional  f - like  sequences . a single nucleotide , either c or t , was added between positions 2296 and 2297 . the theoretical translation of this modified orf yields a protein of 546 aa , the same length as the nipah virus f protein ( data not shown ) . a pairwise alignment of the resulting protein with the f protein of nipah virus shows 32% identity and 53% similarity over 509 aa ( data not shown ) . within this protein , a putative fusion peptide is readily identifiable based on homology to those of other paramyxoviruses ( figure 1b ) . although the cleavage site adjacent to the fusion peptide is typically a basic amino acid , the reported angrem52 cdna sequence has an acidic glutamic acid at this position ( figure 1b ) . both nipah and hendra viruses possess f proteins that are cleaved at the expected site ( figure 1b ) but are apparently processed by novel but ubiquitous proteases . cleavage of these sites can occur even when the residue immediately left of the cleavage site is mutated to a nonbasic residue ( a. maisner , r.e . other features common to paramyxovirus fusion proteins , type i transmembrane glycoproteins , are a signal sequence , a transmembrane domain , and 2-heptad repeats . the 2- heptad repeats play an essential role in membrane fusion and are able to form trimeric coiled coils ( 13 ) . for the putative opmv f , a potential signal sequence from residues 1 to 23 and a potential transmembrane domain is found between residues 497 and 516 . further , the putative opmv f has heptad repeats in the positions expected for a paramyxovirus f protein ( residues 108190 and 428481 ) . the signals that lie between genes include a gene - end signal , an intergenic sequence , and a gene - start signal . the angrem52 sequence , which contains the continuous sequence for the m and f genes , possesses a sequence with similarity to the regulatory sequences in other paramyxoviruses ( figure 1a ) . based on the reported sequence , an orf is present from nt 90 to 1133 , and the theoretical translation of this orf yields a protein with homology to paramyxovirus p proteins but shorter than reported p proteins . however , the insertion of a single t residue between nucleotides at positions 1130 and 1131 results in a single reading frame that ends at position 1579 of the reported angrem104 sequence and encodes a protein of 496 aa . alignment of the modified protein sequence to the nipah virus p protein shows 20.1% amino acid identity over the length of the putative opmv protein . for example , c proteins are encoded by alternate orfs near the 5 end of p genes in a number of paramyxoviruses . in addition , through the process of  rna editing ,  the site - specific insertion of nontemplate encoded nucleotides by the viral polymerase , additional proteins , such as v and w proteins , can be produced ( 13 ) . these latter proteins share amino - terminal sequences with the p proteins but differ after the editing site and thus have distinct carboxy - terminal ends ( 13 ) . in the case of v proteins , the unique carboxy - terminus is characterized by a relatively conserved cysteine - rich domain . analysis of the modified angrem104 sequence identifies a c orf ( from positions 109 to 598 ) potentially encoding a 163-aa protein . in addition , a possible rna editing signal similar to that found in the p gene of other paramyxoviruses is present ( figure 1a ) . one or 2 additional g residues added to the newly synthesized mrna transcribed from this template sequence ( i.e. , the singly edited mrna sequence would then be aaaaaaggg ) would give rise to mrnas encoding a v or w protein ( figure 1c ) . the v orf would begin at nt 90 of our modified angrem104 sequence and end at position 964 of the modified angrem104 sequence ( this numbering does not count the additional g residue found in the edited mrna ) . the carboxy - terminus of the predicted v protein is cysteine - rich , as expected for a paramyxovirus v protein ( figure 1d ) . the w orf would also begin at nt 90 but would end at nt 1027 of the original angrem104 sequence ( again not including the 2 extra g nucleotides introduced by editing [ figure 1c ] ) . based on the similarity of the predicted angrem52 and angrem104 sequences to paramyxovirus protein sequences , a reasonable conclusion is that angrem52 and angrem104 are actually paramyxovirus genes . phylogenetic comparison of the putative opmv p , m , and f proteins suggests that the putative opmv belongs to a previously uncharacterized genus in the paramyxovirus family . comparison of the m proteins may provide the most compelling argument for the uniqueness of the putative opmv , given that an intact orf was present in the angrem52 nucleotide sequence and did not require additional manipulation before analysis . the putative opmv m is found to be slightly more similar to the henipah virus genus than to other paramyxoviruses but distinct from even nipah and hendra virus ( figure 2a ) . likewise , the putative opmv f protein shows the highest degree of sequence identity with the f protein of tupaia paramyxovirus ( figure 2b ) . finally , analysis of the putative opmv p gene places the putative opmv protein on a separate branch with slightly greater similarity to the p proteins of hendra and nipah virus ( figure 2c ) . final evidence for classifying the putative opmv in a distinct phylogenetic group is the fact that the nucleotide sequences of its genes do not show substantial similarity to other paramyxoviruses ( data not shown ) . typically , notable nucleotide identity is seen between members of a paramyxovirus genus but is not seen when nucleotide sequences are compared across genera . for example , morbillivirus m genes share nucleotide identity with one another but not with the m genes of henipah viruses ( data not shown ) . a ) phylogenetic tree showing the relationship of the putative opmv m protein to the m proteins of other paramyxoviruses representative of the various genera in the family paramyxoviridae . b ) phylogenetic tree showing the relationship of the putative opmv f protein to the f proteins of other representative paramyxoviruses . c ) phylogenetic tree showing the relationship of the putative opmv p protein to the p proteins of other representative paramyxoviruses . sequence alignments were made with the clustalw method of the alignx program of the vector nti6 software package . the trees were generated from these alignments by using neighbor - joining methods through the computer program mega version 2.1 ( available from http://www.megasoftware.net/ ) . the position of the putative opmv sequences are indicated by arrows ; distance bars , which represent 0.2 amino acid changes per position , are shown below the trees . the sequences from which the trees were constructed are as follows : mossman , mossman virus ( nc_005339 ) ; tupaia , tupaia paramyxovirus ( nc_002199 ) ; niv , nipah virus ( nc_002728 ) ; hev , hendra virus ( nc_001906 ) ; sev , sendai virus ( ab065188 ) ; hpiv-1 , human parainfluenza virus 1 ( nc_003461 ) ; hpiv-3 , human parainfluenza virus type 3 ( nc_001796 ) ; bpiv-3 , bovine parainfluenza virus type 3 ( af178655 ) ; hrsv , human respiratory syncytial virus ( gi:133665 ) ; brsv , bovine respiratory syncytial virus ( nc_001989 ) ; pmv , pneumonia virus of mice ( ay573814)amnv , avian metapneumovirus ( ay028582 ) ; hmnv , human metapneumovirus ( nc_004148 ) ; sv5 , simian paramyxovirus sv5 ( d13868 ) ; hpiv-2 , human parainfluenza virus type 2 ( nc_003443 ) ; muv , mumps virus ( ay309060 ) ; ndv , newcastle disease virus ( nc_002617 ) ; mev , measles virus ( af266288 ) ; rpv , rinderpest virus ( ab021977 , m21514 , m34018 ) ; dmv , dolphin morbillivirus ( nc_005283 ) ; cdv , canine distemper virus ( nc_005283 ) . polymerase chain reaction ( rt - pcr)based method from primary human mesangial cells ( 10,11 ) . basal expression of each gene was detected , but each gene was identified as an angiotensin ii  induced gene ( 10,11 ) . the apparent presence of viral genes in a primary human cell culture system is intriguing . the presence of these genes could reflect the presence of a virus in any of several states . these include the presence of an actively replicating , fully competent virus ; the presence in the cells of a persistent virus infection ; or the presence of a replicating but defective virus ( 14 ) . although unlikely , these genes could also reflect the integration into the cellular genome of a viral genome as cdna ( 15,16 ) . we have performed several experiments in an effort to determine whether these paramyxoviruslike sequences are universally present within the human genome or whether they represent a very common infection found in human mesangial cells . searches of publicly available human and mouse sequence databases have not identified proteins or predicted proteins with homology to the putative opmv sequences ( data not shown ) . we obtained a human 12-tissue , multiple tissue , northern blot from clontech and probed this with recombinant probes corresponding to the putative opmv m and p genes . no specific signal could be obtained under conditions in which a -actin probe efficiently recognized its mrna ( data not shown ) . we also screened 4 lots of clonetics primary human mesangial cells ( cambrex , east rutherford , nj ) for the presence of the putative opmv mrnas and products when the cells were untreated and after treatment with a range of concentrations of human angiotensin ii ( sigma chemical co. , st . rt - pcr analysis using a number of primer sets for each of the putative opmv genes yielded consistently negative results , and antibodies raised against recombinant forms of the putative opmv m and p proteins failed to detect the putative opmv proteins ( data not shown ) . based on these data , it appears that the putative opmv genes are not human genes and are not universally expressed in human mesangial cells . the source of opmv , the genes of which were identified as angrem52 and angrem104 , remains unclear . infection of the cells after their establishment in culture can not be excluded , nor can contamination of the pcr reactions used to identify angrem52 and angrem104 be ruled out . however , the cells may also have been infected in vivo before the generation of the primary cell culture . in this respect , the putative opmv might be similar to simian virus 5 ( sv5 ) , which can cause persistent infection in monkey kidneys and be detected in monkeys for long periods after initial infection ( 14 ) . given the possibility that the putative opmv infects human cells , the possible association of the putative opmv with human disease is worth exploring .
in public databases , we identified sequences reported as human genes expressed in kidney mesangial cells . 
 the similarity of these genes to paramyxovirus matrix , fusion , and phosphoprotein genes suggests that they are derived from a novel paramyxovirus . 
 these genes are sufficiently unique to suggest the existence of a novel paramyxovirus genus .