Quantitative and Qualitative Involvement of P3N-PIPO in Overcoming Recessive Resistance against Clover Yellow Vein Virus in Pea Carrying the cyv1 Gene

Abstract

In pea carrying cyv1, a recessive gene for resistance to Clover yellow vein virus (ClYVV), ClYVV isolate Cl-no30 was restricted to the initially infected cells, whereas isolate 90-1 Br2 overcame this resistance. We mapped the region responsible for breaking of cyv1-mediated resistance by examining infection of cyv1 pea with chimeric viruses constructed from parts of Cl-no30 and 90-1 Br2. The breaking of resistance was attributed to the P3 cistron, which is known to produce two proteins: P3, from the main open reading frame (ORF), and P3N-PIPO, which has the N-terminal part of P3 fused to amino acids encoded by a small open reading frame (ORF) called PIPO in the ؉2 reading frame. We introduced point mutations that were synonymous with respect to the P3 protein but nonsynonymous with respect to the P3N-PIPO protein, and vice versa, into the chimeric viruses. Infection of plants with these mutant viruses revealed that both P3 and P3N-PIPO were involved in overcoming cyv1-mediated resistance. Moreover, P3N-PIPO quantitatively affected the virulence of Cl-no30 in cyv1 pea. Additional expression in trans of the P3N-PIPO derived from Cl-no30, using White clover mosaic virus as a vector, enabled Cl-no30 to move to systemic leaves in cyv1 pea. Susceptible pea plants infected with chimeric ClYVV possessing the P3 cistron of 90-1 Br2, and which were therefore virulent toward cyv1 pea, accumulated more P3N-PIPO than did those infected with Cl-no30, suggesting that the higher level of P3N-PIPO in infected cells contributed to the breaking of resistance by 90-1 Br2. This is the first report showing that P3N-PIPO is a virulence determinant in plants resistant to a potyvirus.

P3N-PIPO, a Frameshift Product from the P3 Gene, Pleiotropically Determines the Virulence of Clover Yellow Vein Virus in both Resistant and Susceptible Peas

“…Finally, we showed a consistent gradation of virulence among ClYVV isolates in cyv1-carrying peas (recessive resistance) and two susceptible pea lines. This study, combined with data from our previous studies (7), shows that P3N-PIPO is involved in both breaking of recessive resistance and disease expression. We suggest that the pleiotropic effects of P3N-PIPO in determining ClYVV virulence in pea result in a trade-off between the breaking of recessive resistance (cyv1) and maintenance of host viability, which is presumably controlled by a Cyn1-mediated immune response.…”

“…Sequences of the primers used for vector construction are available upon request. The construction of ClYVV chimeric clones Cl-P1HC, Cl-BB, Cl-NS, Cl-SB, Cl-RB, and Cl-RB M28R with GFP (RB/P3 and P3N-PIPO M28R in reference 7) was described previously (7). To make Cl-RB without GFP, the SalI- BamHI fragment of Cl-BB with GFP was used to replace that of Cl-RB with GFP.…”

“…In this study, we constructed pWCl/P3-No. 30 (7). We cloned P3N-PIPO of Cl-No.30 as a fragment from the 5= end of P3 to the position corresponding to the stop codon of Cl-RB PIPO (located downstream of the stop codon of the PIPO frame of Cl-No.30).…”

“…Our previous analysis revealed that P3N-PIPO, which consists of the N-terminal amino acids of P3 followed by a small peptide called PIPO, encoded in the ϩ2 reading frame (8)(9)(10), is a major determinant for breaking of cyv1 resistance (7). We suggested that the higher level of accumulation of P3N-PIPO in Cl-90-1 Br2 than in Cl-No.30 contributes to the breaking of resistance (7). P3N-PIPO has an essential role in virus cell-to-cell movement (10).…”

The P1N-PISPO trans -Frame Gene of Sweet Potato Feathery Mottle Potyvirus Is Produced during Virus Infection and Functions as an RNA Silencing Suppressor

“…Since the PIPO coding sequence was first described in potyviruses, the new gene product P3N-PIPO has attracted much interest, leading to the identification of several important associated functions (8)(9)(10)(11)(12). However, the mechanism by which P3N-PIPO is produced remained unclear until recently, when we and another team independently found evidence that PIPO is expressed by a polymerase slippage mechanism (13,14).…”