Control of Oriented Tissue Growth through Repression of Organ Boundary Genes Promotes Stem Morphogenesis

Abstract

SummaryThe origin of the stem is a major but poorly understood aspect of plant development, partly because the stem initiates in a relatively inaccessible region of the shoot apical meristem called the rib zone (RZ). We developed quantitative 3D image analysis and clonal analysis tools, which revealed that the Arabidopsis homeodomain protein REPLUMLESS (RPL) establishes distinct patterns of oriented cell division and growth in the central and peripheral regions of the RZ. A genome-wide screen for target genes connected RPL directly to many of the key shoot development pathways, including the development of organ boundaries; accordingly, mutation of the organ boundary gene LIGHT-SENSITIVE HYPOCOTYL 4 restored RZ function and stem growth in the rpl mutant. Our work opens the way to study a developmental process of importance to crop improvement and highlights how apparently simple changes in 3D organ growth can reflect more complex internal changes in oriented cell activities.

Auxin-Induced Modulation of ETTIN Activity Orchestrates Gene Expression in Arabidopsis

“…ETT expression in the meristem followed a phyllotactic pattern coinciding with the region where new floral primordia were emerging ( Figure 7A) and overlapped with previously reported RPL expression in this region ( Figure 7A; Bencivenga et al, 2016). Interestingly ett-3, rpl-2, ett-3 rpl-2, and ETT C2S mutant plants (F) Heat map generated with Genevestigator showing expression of the 77-set and 59-set targets (which combined create the 136-set targets).…”

“…The mutant line ett-3, pETT:ETT-GFP complementation line, and pETT:ETT C2S transgenic line (Simonini et al, 2016); the pRPL:RPL-GFP complementation line (Bencivenga et al, 2016); and the DR5:GFP, DII-VENUS, pPIN1:PIN1-GFP, pPIN3:PIN3-GFP, pASB1:GUS, and pTAA1:GFP-TAA1 lines (Benková et al, 2003;Brunoud et al, 2012;Zádníková et al, 2010;Stepanova et al, 2005Stepanova et al, , 2008 have been previously described. We obtained the following T-DNA insertional lines from the SALK collection (Alonso et al, 2003): AT1G22490 (bHLH094/ TEC1), SALK_109362C; AT2G28080 (TEC3), SALK041660 (tec3-1) and SALK_139804 (tec3-2); AT2G42300 (bHLH048/TEC4), SALK_135489C and SALK_013047C; AT3G57800 (bHLH060/TEC5), SALK_134005C; and AT1G72210 (bHLH096/TEC6), SALK_060037C.…”

“…ChIP-seq and transcriptome data sets for a number of ETT partners are already available in the public domain (Bencivenga et al, 2016;Santuari et al, 2016) and comparative studies are therefore possible to identify common targets and unravel modes of regulation. Here, we compared the list of 77 target genes bound and regulated by ETT in an IAA-dependent manner with the target gene data set of the ETT-interacting REPLUMLESS (RPL) protein (Bencivenga et al, 2016) ( Figure 5A). The RPL gene encodes a homeobox transcription factor that was chosen because it, like ETT, has a prominent role in reproductive tissue development (Roeder et al, 2003).…”

“…Out of a list of 24 targets shared between ETT and RPL, we identified seven where strong binding was detected in the same region ( Figure 5A; Supplemental Data Set 1). Out of these seven, we decided to focus on four genes ( Figure 5A), which according to the Bio Analytical Resource for Plant Biology (BAR; http://bar.utoronto.ca/) are expressed in meristematic and carpeloid tissues where both ETT and RPL are expressed (Sessions et al, 1997;Roeder et al, 2003;Bao et al, 2004;Bencivenga et al, 2016;Simonini et al, 2016). The set of four ETT-RPL IAA-dependent common target genes included YPT/RAB GTPASE INTERACTING PROTEIN 4B (YIP4B) and SMAX1-LIKE5 (SMXL5), which are involved in secretion of cell wall polysaccharides and strigolactone signaling (Gendre et al, 2013;Wallner et al, 2017), respectively.…”

“…ChIP was performed in triplicate as previously described (Simonini et al, 2016;Bencivenga et al, 2016) using MultiMACS GFP isolation kits, pETT: ETT-GFP in ett-3 line, and wild-type plants (negative control). As with RNA-seq, the material used included meristem and young flowers up to and including stage 10 (Smyth et al, 1990).…”