The Roles of Phosphorylation and SHAGGY-Like Protein Kinases in Geminivirus C4 Protein Induced Hyperplasia

Abstract

Even though plant cells are highly plastic, plants only develop hyperplasia under very specific abiotic and biotic stresses, such as when exposed to pathogens like Beet curly top virus (BCTV). The C4 protein of BCTV is sufficient to induce hyperplasia and alter Arabidopsis development. It was previously shown that C4 interacts with two Arabidopsis Shaggy-like protein kinases, AtSK21 and 23, which are negative regulators of brassinosteroid (BR) hormone signaling. Here we show that the C4 protein interacts with five additional AtSK family members. Bikinin, a competitive inhibitor of the seven AtSK family members that interact with C4, induced hyperplasia similar to that induced by the C4 protein. The Ser49 residue of C4 was found to be critical for C4 function, since: 1) mutagenesis of Ser49 to Ala abolished the C4-induced phenotype, abolished C4/AtSK interactions, and resulted in a mutant protein that failed to induce changes in the BR signaling pathway; 2) Ser49 is phosphorylated in planta; and 3) plant-encoded AtSKs must be catalytically active to interact with C4. A C4 N-myristoylation site mutant that does not localize to the plasma membrane and does not induce a phenotype, retained the ability to bind AtSKs. Taken together, these results suggest that plasma membrane associated C4 interacts with and co-opts multiple AtSKs to promote its own phosphorylation and activation to subsequently compromise cell cycle control.

Nucleocytoplasmic Shuttling of Geminivirus C4 Protein Mediated by Phosphorylation and Myristoylation Is Critical for Viral Pathogenicity

“…Previous studies have predicted that the C4 proteins of several geminiviruses could be modified with a myristoyl lipid anchor (14-carbon saturated fatty alkyl chain) on the N-terminal glycine residue, and that myristoylation is essential for the induction of symptoms (Fondong et al, 2007;Piroux et al, 2007;Dogra et al, 2009;Mills-Lujan and Deom, 2010;Mills-Lujan et al, 2015;Hipp et al, 2016;Rosas-Diaz et al, 2018). Additionally, Mills-Lujan et al (2015) reported that phosphorylation of BCTV C4 by AtSKh/BIN2 was important for symptom development. Hipp et al (2016) detected phosphorylation and myristoylation of AC4 from African cassava mosaic virus in yeast using mass spectrometry.…”

Geminiviruses and Plant Hosts: A Closer Examination of the Molecular Arms Race

“…Global transcriptomic changes upon monopartite TYLCSV infection in tomato showed the up-regulation of genes involved in phytohormone metabolism, nucleic acid metabolism and ubiquitin-autophagy pathway, whereas the down-regulated genes were involved in primary metabolism, methylation-dependent chromatin silencing [ 123 ]. Interactions of curtovirus derived C4 and Arabidopsis thaliana SHAGGY-like protein kinases (AtSKs) were implicated in hyperplasia observed during BCTV infection [ 124 ].…”

The Ever-Expanding Role of C4/AC4 in Geminivirus Infection: Punching above Its Weight?

“…One of the functional characteristics of C4/AC4 is its induction of hyperplasia and tumorigenic growth in infected plant tissues (Mills-Lujan et al, 2015). This property is consistent with the fact that viruses commonly manipulate the cell cycle to create cellular conditions favorable for viral genome replication by encoding proteins that target host cell cycle regulators.…”

“…Another key function of C4/AC4 related to cell cycle regulation is the induction of characteristic viral symptoms. For instance, East African cassava mosaic Cameroon virus (EACMCV) was shown to cause crinkling symptoms (Fondong et al, 2007) and Beet curly top virus (BCTV) C4 causes hyperplasia symptoms similar to symptoms observed during a BCTV infection (Mills-Lujan et al, 2015). This is presumed to be a consequence of C4/AC4 interaction with CLAVATA 1 (CLV1) receptor kinase that functions in meristem maintenance .…”