A tomato prolyl-4-hydroxylase causes relocation of abscission zone and alters abscission kinetics

Abstract

The detachment of organs is controlled by highly regulated molecular mechanisms. The position of the tomato abscission zone (AZ) is defined by the ratio of the proximal to distal part of the pedicel. In this study, the ratio was altered due to a shift in the position of the AZ which was attributed to shorter and longer pedicels of SlP4H3 RNAi and OEX lines due to changes on cell division and expansion in AZ and distal part. This might be associated with LM2- and JIM8-AGPs which increased in OEX and decreased in RNAi lines throughout the pedicel. The JIM13 AGPs were downregulated in the flower AZ of OEX lines, pointing to a role on abscission regulation. In addition, Co-IP in flower AZ with SlP4H3-GFP fusion proteins showed interaction with LM2-, JIM13- and JIM8-epitopes suggesting proline hydroxylation by SlP4H3. The lower content of methyl-esterified HGs and higher of demethyl-esterified HGs in the AZs of RNAi lines might be responsible for increased rigidity of the AZ cell walls, accounting for the higher force required for AZ tissue detachment to occur. Moreover, ethylene-induced flower abscission was accelerated in the RNAi lines and delayed in OEX lines, while exactly the opposite response was observed in the red ripe fruit AZs. This was partly attributed to alterations in the expression of cell wall hydrolases. Overall, these results indicate that P4Hs might regulate molecular and structural features of cell walls in the AZ as well as abscission progression by regulating the structure and function of AGPs.

The modified activity of prolyl 4 hydroxylases reveals the effect of arabinogalactan proteins on changes in the cell wall during the tomato ripening process

“…Two SlP4H3 RNAi independent lines, #1 and #7, and two SlP4H3 overexpression lines, OEX#1, OEX#2 were previously produced ( Perrakis et al., 2019 , 2021 ). Moreover, two additional, independent overexpression lines were created, cGFP OEX (cGFP) and nGFP OEX (nGFP), comprising a construct in which a GFP tag was either at the C- or the N-terminus of the SlP4H3 cDNA under the control of the 35S promoter ( Perrakis et al., 2021 ). The expression levels of SlP4H3 in the two silencing and the four overexpression lines were determined at the four ripening stages ( Figure 1 ).…”

“…Homozygous T2 and T3 generation tomato ( Solanum lycopersicum cv. ‘Ailsa Craig’) plants of two RNAi, RNAi#1, RNAi#7, and four overexpression, OEX#1, OEX#2, and cGFP OEX (cGFP), nGFP OEX (nGFP), lines were prepared ( Perrakis et al., 2021 ). Plants of the T2 and T3 tomato lines were grown under hydroponic conditions in perlite sacks in one glasshouse at Chania, Greece under standard conditions of temperatures at the range of 18-27°C and relative humidity at the range of 60-80% humidity for approximately one year.…”

“…In three stable RNAi transgenic lines of SlP4H3 , abscission was delayed in overripe tomato fruits indicating involvement in the progression of this developmental process ( Perrakis et al., 2019 ). Moreover, suppression and overexpression of SlP4H3 in stable tomato transgenic lines resulted in changes in the content of AGPs-bound epitopes in flower abscission zones and/or distal and proximal tissue ( Perrakis et al., 2021 ). In addition, Arabidopsis mutants of the β-linked glucuronic acid transferases resulted in lower content of glucuronic acids on AGPs leading to severe developmental phenotypes such as limited seedling growth and sterility which were associated with perturbed calcium waves indicating reduced binding of calcium due to insufficient glucuronidation of AGPs ( Lopez-Hernandez et al., 2020 ).…”

The modified activity of prolyl 4 hydroxylases (P4Hs) reveals the effect of arabinogalactan proteins (AGPs) on changes in the cell wall during the tomato ripening process

“…Homozygous T2 and T3 generation tomato (Solanum lycopersicum cv. 'Ailsa Craig') plants of two RNAi, RNA#1, RNA#7, and four overexpression, OEX#1, OEX#2, and cGFP OEX (cGFP), nGFP OEX (nGFP), lines were grown in the glasshouses at Chania, Greece under standard conditions (Perrakis et al, 2021). Tomato fruits were harvested at the Breaker (BR), Turning (TU), Pink (PINK), and Red Ripe (RR) previously described by 5 Nakatsuka and coworkers (1998).…”

“…Two SlP4H3 RNAi independent lines, #1 and #7, and two SlP4H3 overexpression lines, OEX#1, OEX#2 were previously produced (Perrakis et al, 2019(Perrakis et al, , 2021. Moreover, two additional, independent overexpression lines were created, cGFP OEX (cGFP) and nGFP OEX (nGFP), comprising a construct in which a GFP tag was either at the C-or the N-terminus of the SlP4H3 cDNA under the control of the 35S promoter (Perrakis et al, 2021). The expression levels of SlP4H3 in the two silencing and the four overexpression lines were determined at the four ripening stages (Figure 1).…”