Betaine Accumulation andI'4ClFormate Metabolism inWater- stressed Barley Leaves" 2

Barley (Hordeum vulgare L.)plants atthethree-leaf stage werewaterstressed byflooding therooting medium withpolyethylene glycol 6000with anosmotic potential of-19bars, orbywithholding water. While leaf water potential feli andleaf kill progressed, thebetaine (trimethylgycine) content ofthesecond leaf blade rosefromabout 0.4micromole toabout1.5 micromoles in4days. Thetimecourse ofbetaine accumulation resembled thatofproUne accumulation. Choline levels inunstressed second leaf blades werelow(<0.1 micromole perblade) andremained lowduring water stress. Uponrelef ofstress, betaine-like proline-remained atahigh concentration indrought-killed leaf zones, butbetaine didnotdisappear as rapidly asproline fromviable leaf tissue during recovery. WhenImethyl-14Clcboline wasapplied tosecond leaf blades ofintact plants inthegrowth chamber, water-stressed plants metabolized 5to10 times more'Clabel tobetaine thancontrol plants during 22hours. When infiltrated with tracer quantities of114Clformate andincubated forvarious times indarkness orlight, segments cutfromwater-stressed leaf blades incorporated about 2-to10-fold more14Cinto betaine thandidsegments fromunstressed leaves. Insegments fromstressed leaves incubated with I14CIformate forabout18hoursindarkness, betaine wasalways the principal 14C-labeled soluble metabolite. This14Clabel waslocated exclusively intheN-methyl groups ofbetaine, demonstrating thatreducing equivalents wereavailable instressed leaves forthereductive steps of methyl group biosynthesis fromformate. Incorporation of"Cfromformate into choline wasalso increased instressed leaf tissue, butcholine wasnot amajor product formed from114Clformate. These results areconsistent with anetdenovosynthesis ofbetaine from 1-and2-carbon precursors during waterstress, andindicate thatthe betaine soaccumulated maybeametabolically inert endproduct. contribute tocontinued growth orsurvival during stress) andthose whicharedeleterious ormerely incidental consequences ofthe stress. These distinctions mustbedrawnbefore metabolic criteria canbeusedinplant-breeding programs targeted atimproving stress resistance. Aspartofaprogram toinvestigate potential metabolic criteria forbreeding plants adapted towater stress (11), weherepresent dataonbetaine andproline accumulation in water-stressed barley leaves. Betaine accumulation appears promising asametabolic index ofcumulative waterstress experience.