Abstract
Aim: The transport of sugars into grape berry mesocarp cells across the plasma and vacuolar membranes after onset of ripening is a complex process. Elements of the sugar transport mechanism may be assessed by exposing the mesocarp cells and investigating sugar movement across the membranes. The purpose of this study was to gain insights into the nature of the transport mechanism by creating conditions conducive to hexose efflux from the peeled berry.Methods and results: The experimental technique employed was a derivate of the ‘berry-cup’ technique. The skin of ripening cv. Shiraz berries was peeled in situ and, after an initial wash, hexose efflux into a collection medium (MES buffer) was monitored. Additionally, during the period of intensive sugar accumulation (one week after veraison) and two weeks later, hexose efflux was assessed following three modifications: (i) using berries excised from the vine, (ii) using MES buffer (2-(N-morpholino)ethanesulfonic acid, pH 5.5) containing PCMBS (p-chloromercuribenzenesulfonic acid, 1mM), and (iii) using cold (10°C) or warm (40°C) MES buffer. Hexose quantities collected into the buffer were dependent on ripening stage and buffer temperature, but they were not dependent on an intact berry-to-cluster connection. The inhibitory effect of PCMBS was observed early in ripening, but not two weeks later.Conclusions: These results lead us to the conclusion that the origin of the collected hexoses was vacuolar as opposed to vascular, and that the hexose efflux mechanism is differently sensitive to PCMBS at the two stages of ripening.Significance and impact of the study: This simple technique was effective at providing insights into hexose transport within the grape berry at the cellular level.
Highlights
The accumulation of glucose and fructose into thevacuolesofgrapeberrymesocarpcellsisone of the most integral but complex processes of berry ripening
Sugaraccumulationintothegrapeberryis essentialtowinequality.Aftertheonsetofgrape berry ripening, phloem unloading follows an apoplasmicrouteintothemesocarptissue (Zhanget al.,2006).Intheapoplast,mostofthe unloaded sucrose is cleaved by cell wall invertases,andimportedintothecellsasglucose and fructose
During the same 5-week experimental period, glucose and fructose release from peeled berries into the buffer solutions increased with berry ripeness (Figure 3)
Summary
The accumulation of glucose and fructose into thevacuolesofgrapeberrymesocarpcellsisone of the most integral but complex processes of berry ripening. This process serves as an important mechanism for solute potential regulation (Wada et al, 2008) and it turns the fruit into a tasty seed-dispersal mechanism mediated through birds. Sugaraccumulationintothegrapeberryis essentialtowinequality.Aftertheonsetofgrape berry ripening, phloem unloading follows an apoplasmicrouteintothemesocarptissue (Zhanget al.,2006).Intheapoplast,mostofthe unloaded sucrose is cleaved by cell wall invertases,andimportedintothecellsasglucose and fructose. Several membrane proteins have beenidentifiedastakingpartinthesugar transportmechanism,andsomeofthem(sucrose transporters and SWEET family of 46 sugar transporters)mayperformsugartransportinboth directions across the membrane (reviewed by Lecourieuxet al.,2014).Thegrapevinegenome probably contains 20 putative hexose transportersbutjustafewofthesehavea significant role in berry hexose accumulation (Fillion et al, 1999 ; Vignault et al, 2005 ; Zhanget al.,2008 ;Afoufa-Bastienet al.,2010)
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