Further Evidence that Viroplasms Are the Site of Cauliflower Mosaic Virus Genome Replication by Reverse Transcription during Viral Infection

Abstract

SUMMARYCauliflower mosaic virus (CaMV) is a DNA plant virus that replicates its genome through an RNA intermediate. The cytoplasmic step of CaMV DNA replication was studied using a fraction consisting of purified viroplasms, which are virus-specific inclusion bodies accumulating in the infected plant cells. The isolated viroplasms retain a DNA polymerase activity able to synthesize CaMV DNA from endogenous templates. A further characterization of the viral DNA sequences produced in the isolated inclusion bodies indicates that newly synthesized DNA, mostly of polarity opposite to that of viral RNA, is single-stranded and partly associated with RNA by base-pairing. In addition to an RNA-dependent DNA polymerase activity, RNA molecules, which presumably originate from the viral RNA template used for reverse transcription, are found to accumulate in the purified inclusion bodies. Furthermore, a small DNA molecule strongly labelled in the purified fraction has been characterized and corresponds to the CaMV reverse transcription intermediate sa-DNA. These results provide further evidence that the reverse transcription of CaMV RNA occurs in the viroplasms. Additional data are presented which suggest that CaMV replication could occur in virion-related particles.

Cauliflower mosaic virus: still in the news

“…However, virus‐like particles were rarely formed, if any, suggesting that CaMV capsid assembly might depend on additional factors. One of them could be CaMV P6 protein which was shown to interact with the basic domain of P4 and may thus represent a scaffolding protein (Himmelbach et al ., 1996), especially since both replication of the viral genome and its encapsidation occur on the P6‐formed inclusion bodies (Marsh and Guilfoyle, 1987; Mazzolini et al ., 1989). These observations, and the fact that capsid proteins interact with the pre‐genomic 35S RNA strongly suggest that genome replication and morphogenesis are coupled as already described for Hepadnaviruses.…”

The Open Reading Frame VI Product ofCauliflower mosaic virusIs a Nucleocytoplasmic Protein: Its N Terminus Mediates Its Nuclear Export and Formation of Electron-Dense Viroplasms

“…Electron-dense viroplasms are a hallmark of infection of plant cells by caulimoviruses and soymoviruses. They probably play an important role in the infectious cycle because they are the sites of protein synthesis, viral genome replication, morphogenesis, and storage of the newly formed virions (Mazzolini et al, 1989;Rothnie et al, 1994). Other CaMV proteins have been detected in the electron-dense viroplasms (Drucker et al, 2002), but none of them seems to be required for their formation because transgenic Arabidopsis plants expressing P6 alone contain inclusion body-like structures (Cecchini et al, 1997).…”

P6 protein of Cauliflower mosaic virus, a translation reinitiator, interacts with ribosomal protein L13 from Arabidopsis thaliana

“…In particular it is involved in host-range determination (Schoelz & Shepherd, 1988), symptomatology (Daubert & Routh, 1990;Wintermantel et al, 1993;Broglio, 1995), and probably viral morphogenesis (Himmelbach et al, 1996). Moreover, P6 is also the major component of the membrane-free cytoplasmic inclusion bodies called viroplasms (Xiong et al, 1982), where the main steps of the replication cycle occur (Mazzolini et al, 1989).…”