Proteomic analysis of xylem vessel cell differentiation in VND7-inducible tobacco BY-2 cells by two-dimensional gel electrophoresis

Abstract

The xylem vessel is an essential structure for water conduction in vascular plants. Xylem vessel cells deposit thick secondary cell walls and undergo programmed cell death, to function as water-conducting elements. Since the discovery of the plant-specific NAC domain-type VASCULAR-RELATED NAC-DOMAIN (VND) transcription factors, which function as master switches of xylem vessel cell differentiation in Arabidopsis, much has been learned about the transcriptional regulatory network of xylem vessel cell differentiation. However, little is known about proteome dynamics during xylem vessel cell differentiation. Here, we performed two-dimensional electrophoresis-based proteomic analysis of xylem vessel cell differentiation using a transgenic tobacco BY-2 cell line carrying the VND7-inducible system (BY-2/35S::VND7-VP16-GR), in which synchronous trans-differentiation into xylem vessel cells can be induced by the application of a glucocorticoid. Of the 47 spots revealed by gel electrophoresis, we successfully identified 40 proteins. Seventeen proteins, including several well-characterized proteins such as a cysteine protease and serine carboxypeptidase (involved in programmed cell death), were upregulated after 24 h of induction. However, previous transcriptomic analysis showed that only eight of these proteins are upregulated at the transcriptional level during xylem vessel cell differentiation in BY-2/35S::VND7-VP16-GR cells. These findings suggest that post-transcriptional regulation strongly affects proteomic dynamics during xylem vessel cell differentiation.

Modeling cross-regulatory influences on monolignol transcripts and proteins under single and combinatorial gene knockdowns in Populus trichocarpa

“…In addition to changes in transcript abundance, we also observed several cases where 478 monolignol protein abundances were significantly altered when their transcripts were 479 not. This behavior has previously been observed in secondary cell wall proteins of 480 Arabidopsis [37,38] and in tobacco during cell differentiation [39]. Compared to 481 transcriptional regulation, less is known about the role of post-transcriptional and 482 post-translational regulatory mechanisms on monolignol biosynthesis.…”

“…In addition to transcriptional regulation, post-transcriptional and post-translational 49 regulatory mechanisms relating to translation or protein degradation can play a critical 50 role in protein abundance [13,35,36]. Differences in expression of transcripts and 51 proteins suggesting such regulatory mechanisms have been found for genes encoding cell 52 wall proteins in Arabidopsis thaliana [37,38] and for genes involved in tobacco xylem cell 53 differentiation [39]. These mechanisms were also proposed to explain the poor 54 correlations between some of the monologinol gene transcripts and proteins in some 55 transgenic P. trichocarpa [4].…”

Creating vessel elements in vitro: Towards a comprehensive understanding of the molecular basis of xylem vessel element differentiation

“…80-90%) and have been used to determine the transcriptional networks downstream of VND6 and VND7 (Ohashi- Yamaguchi et al 2011;Zhong et al 2010). Moreover, the VND7inducible system has revealed many novel cellular and molecular mechanisms involved in xylem vessel element differentiation (Endo et al 2015;Goué et al 2013;Kawabe et al 2018;Li et al 2016;Noguchi et al 2018;Ohtani et al 2016Ohtani et al , 2018Schuetz et al 2014;Takenaka et al 2018;Watanabe et al 2015Watanabe et al , 2018.…”

“…Using these systems, homogenous cells at the same stage of differentiation can be collected and analyzed, facilitating the identification of novel factors involved in specific stages of cell differentiation. Indeed, in vitro induction systems have already revealed many regulatory aspects of xylem vessel element differentiation, including novel key regulators of xylem vessel cell differentiation and the stage-specific regulation of the transcriptome, proteome, and metabolome (Demura et al 2002;Endo et al 2015;Fukuda 1997Fukuda , 2004Goué et al 2013;Ito et al 2006;Kawabe et al 2018;Kondo et al 2014Kondo et al , 2015Kondo et al , 2016Kubo et al 2005;Li et al 2016;Motose et al 2004; Copyright © 2019 The Japanese Society for Plant Cell and Molecular Biology Noguchi et al 2018;Fukuda 2012, 2013;Oda et al 2010;Ohtani et al 2016Ohtani et al , 2018Pesquet et al 2010;Schuetz et al 2014;Takenaka et al 2018;Tan et al 2018;Watanabe et al 2015Watanabe et al , 2018.…”

Modeling cross-regulatory influences on monolignol transcripts and proteins under single and combinatorial gene knockdowns in Populus trichocarpa

“…Wang et al, made the simplifying assumption that the abundance of each protein was dependent only on the transcript abundance of its monolignol gene. This simplification does not take into consideration potential epistatic transcriptional, post-transcriptional, or post-translational regulation mechanisms [12,15,16,[19][20][21] that may explain the sometime poor correlations between monolignol gene transcripts and proteins in some transgenic P. trichocarpa [4]. The development of approaches that can more accurately predict how the abundances of interconnected transcripts and proteins change under single and combinatorial transgenic knockdowns, while also providing insight on the topological structure of these cross-influences, would greatly improve our ability to predict how metabolic pathways and phenotypic traits will be altered under these modifications.…”