Pokeweed (Phytolacca americana L.) antiviral protein inhibits Zucchini yellow mosaic virus infection in a dose-dependent manner in squash plants

Abstract

Pokeweed antiviral protein (PAP) of Phytolacca americana L. (pokeweed) is a single-chain ribosome-inactivating protein (RIP) characterized by its ability to depurinate plant ribosomes. Here, we isolated, cloned, and expressed the ribosome inactivating protein (RIP) gene, designated as pokeweed antiviral protein type 1 (PAP I), from the summer leaves of pokeweed collected from the Black Sea region (Turkey). Our findings presented here provide direct evidence that exogenous application of PAP I causes concentration-dependent inhibition of Zucchini yellow mosaic virus (ZYMV) infection on squash plants. Squash plants were exposed to PAP I protein with and without DMSO for four consecutive days. Regular spraying of approximately 30 kDa recombinant PAP I at 2 µg mL -1 concentration prevented treated plants from mechanical virus infection. PAP I showed antiviral activity in 9 plants out of 15 inoculated plants. Remarkably, simultaneous application of PAP, DMSO, and ZYMV did not prevent virus infection, suggesting that PAP did not have any effect on viral RNA. In the absence of ZYMV the purified peptide was not cytotoxic for squash plants, although a reduction of plant size, possibly caused by host ribosome depurination, was observed.

The Plant Ribosome-Inactivating Proteins Play Important Roles in Defense against Pathogens and Insect Pest Attacks

“…Further studies suggested that the exogenous application of PAP appears to enhance the systemic resistance of N. benthamiana to TMV infection ( Zhu et al, 2016 ). Application of PAP I also causes concentration-dependent inhibition of zucchini yellow mosaic virus (ZYMV) infection on squash plants ( Sipahioğlu et al, 2017 ). Dianthins isolated from the leaves of Dianthus caryophyllus mixed with TMV strongly decreased the amount of local lesions on plant leaves of Nicotiana glutinosa ( Stirpe et al, 1981 ).…”

Antiviral Activity of Ribosome-Inactivating Proteins

“…The evaluation of antiviral activity has been done by counting the number of lesions [ 88 , 93 ], the time of onset of symptoms [ 77 , 79 ], the number of infected plants [ 105 ], or the severity of the infection symptoms [ 78 , 115 ]. Virus levels have also been estimated by ELISA [ 99 ], Western blotting analysis [ 81 ], RT-PCR analysis [ 101 ], quantitative real-time PCR analysis [ 81 , 82 ], electron microscopy [ 92 ], or by determining the infection capacity of an extract from the infected plant [ 92 ]. Another approach has been the construction of virus-resistant transgenic plants [ 80 , 102 ].…”

Ribozom İnaktive Eden Proteinlerin Bitki Virüs Hastalıklarının Kontrolünde Kullanılma Olanakları

“…Çizelge 1 Tip1 Ribozom inaktive eden proteinlere örnekler (Hong et al, 1996;Balasaraswathi et al, 1998;Stirpe, 2004;Picard et al, 2005;Sipahioğlu et al, 2017…”

“…Aynı yöntemle üretilen ve saflaştırılan PAP I proteini, kabak bitkisi yapraklarına harici olarak düzenli biçimde uygulandığında 2 μg/ml konsantrasyonunun Zucchini yellow mosaic virus (ZYMV) infeksiyonunu engellediği görülmüştür. Tek başına uygulandığında, purifiye PAP I'in aynı zamanda bitkilerde cüceleşmeye neden olduğu rapor edilmiştir (Sipahioğlu et al, 2017).…”