investigating-interactions-between-histone-modifying-enzymes

Investigating Interactions Between Histone Modifying Enzymes and Transcription Factors <em>in vivo</em> by Fluorescence Resonance Energy Transfer

Epigenetic regulation of gene expression is commonly affected by histone modifying enzymes (HMEs) that generate heterochromatic or euchromatic histone ...

State University of New York

We have developed an efficient method to rapidly generate infectious inoculum of a plant RNA virus and confirmed its infectivity by mechanical inoculation. The method takes advantage of overlap PCR to bypass the cloning steps, which makes it relatively simple, rapid, and inexpensive compared to the traditional methods. Using this approach, inoculum of a tobamovirus, Turnip vein clearing virus (TVCV), was generated. PCR products specific for the 35S promoter and TVCV genome were used as templates for overlap PCR to form a single product containing the full-length TVCV cDNA under the control of the double 35S promoter, and the entire process took only 8 h. This inoculum was infectious in Nicotiana benthamiana, and its infectivity was ca. 67% compared to 0% and 100% with negative and positive controls, respectively. Thus, this rapid method generates efficient infectious inoculum for a plant RNA virus.

Rapid generation of inoculum of a plant RNA virus using overlap PCR.

Cell-to-cell movement is an important step for initiation and spreading of virus infection in plants. This process occurs through the intercellular connections, termed plasmodesmata (PD), and is usually mediated by one or more virus-encoded movement proteins (MP) which interact with multiple cellular factors, among them protein kinases that usually have negative effects on MP function and virus movement. In this study, we report physical and functional interaction between MP of Tobacco mosaic virus (TMV), the paradigm of PD-moving proteins, and a receptor-like kinase BAM1 from Arabidopsis and its homolog from Nicotiana benthamiana. The interacting proteins accumulated in the PD regions, colocalizing with a PD marker. Reversed genetics experiments, using BAM1 gain-of-function and loss-of-function plants, indicated that BAM1 is required for efficient spread and accumulation the virus during initial stages of infection of both plant species by TMV. Furthermore, BAM1 was also required for the efficient cell-to-cell movement of TMV MP, suggesting that BAM1 interacts with TMV MP to support early movement of the virus. Interestingly, this role of BAM1 in viral movement did not require its protein kinase activity. Thus, we propose that association of BAM1 with TMV MP at PD facilitates the MP transport through PD, which, in turn, enhances the spread of the viral infection.

Phu Tri Tran

Department of Biochemistry and Cell Biology

State University of New York

Investigating Interactions Between Histone Modifying Enzymes and Transcription Factors in vivo by Fluorescence Resonance Energy Transfer

Phu Tri Tran

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Biochemistry and Cell Biology

State University of New York

Investigating Interactions Between Histone Modifying Enzymes and Transcription Factors in vivo by Fluorescence Resonance Energy Transfer

Department of Biochemistry and Cell Biology