VII. GENERAL DISCUSSION
This study was conducted in the line of crop improvement. The demand of food supply is increasing along with the growth of human population, and where
inversely, the quality of the environment is declining. Water, in particular, is becoming scarce, salt concentrations along with other pollutants are increasing at
a time when crop production will be expanded to drier regions to produce biomass for food, energy or other purposes. Trehalose is well-known stress
protecting sugar found to accumulate in organisms exposed to stressful conditions. Introducing genes encode trehalose metabolizing enzymes has been
one of research interest. Plethoric phenotype is resulted from the accumulation of T6P when OtsA and or OtsB; TPS1 and or TPP are introduced to plant, e.g. on
tobacco Goddijn et al. 1997: Pilon-Smith et al. 1998, and Welin et al. 2001. Introducing bifunctional fusion of E. coli OtsA-OtsB same as TPS1-TPP
overcome the problem as reported by Garg et al. 2002 and Jang et al. 2003, since T6P produced by OtsA is directly converted to trehalose.
This study assessed an alternative of gene encoding trehalose metabolizing enzyme for such purpose. This pathway involves a single gene only
to produce trehalose from maltose without producing T6P, hoping that unexpected phenotypical defect might be prevented. TRES as trehalose
synthesizing at the same time as trehalose degrading enzyme might be beneficial to the transgenic plant, by which metabolizable sugar is still available even if
trehalose level reach the equilibrium. This proved with the capability of transgenic plants survive on high level trehalose eg 125 mM. As it shown in Chapter 6, that
Arabidopsis bearing TreS grow normally and fertile. The plants were resistant to
trehalose that might be as one of indication withstands stresses including in respone to trehalose-producing pathogens.
Research has now shown that metabolism of trehalose in plants is critical to engineer drought stress resistant plants. It is clear that the presence of
trehalose synthesizing gene improve plant responses toward stresses Garg et al 2002; Jang et al 2003; Zhang et al 2005; Zhang et al 2006. In this experiment
using Arabidopsis in planta by with holding water showed that 30 of transgenic plants recover from drought stress and grow normal after rehydrated.
Excised leaf tests with parameters of leaf water retention, leaf total water lost, leaf dry weight; and inflorescent trehalose content and inflorescent TRES activity
followed by a cluster analysis based on the characters suggests that 33 of the transgenic plants resistant to drought. From leaf water recovery test, it was
observed 20 of leaves recover after 16h keeping on RT. Hence it might be suggested that there is possibility about 30 to obtain transgenic plant with
improved drought response by introducing trehalose synthase gene from Thermobifida fusca
. It was suggested that the improved response toward stress of
transgenic plant bearing trehalose metabolizing genes was due to the increase of soluble carbohydrate as result of the increase of photosynthetic
capacity Garg et al 2002. While Jang et al 2003 suggested, trehalose give global protection against abiotic stresses, yet, they did not suggest the
mechanism. While Avonce et al. 2004 suggested that AtTPS1 is regulator of glucose, abcisic acid, and stress signaling. In this experiment, the capability of
some TfTreS expressing lines withstand drought might also has been implicated with the increase of T6P. Arabidopsis expressing AtTPS1 is sugar and ABA
insensitive, and T6P regulate the expression of ABI4. ABI4 is down regulated by T6P when glucose is presence and is up regulated in the absence of
metabolizable sugar. T6P affect growth and development Schluepmann et al 2004, mutant of AtTPS1 is embryo lethal. Hence, trehalose metabolism is
indispensable for growth Easmond et la. 2002; Schluepmann et al. 2004. This effect on growth was also shown on M. smegmatis Woodruff et al 2007. The
presence of T6P, however, leads to induction of APL3 hence starch is accumulated in chloroplast and growth is arrested. Feeding trehalose to
seedlings increased T6P level Schluepmann et al. 2004. Expressing TfTreS that increase trehalose level, might increase increase T6P level on plant in some
extend, as starch accumulation was proved occurred to some lines Figure 33. The increase, of T6P however, did not retard growth that might be due to the
reverse TfTreS activity that still provides sugar for energy source. Furthermore, the presence of the sugar down regulates ABI4, hence ABA production is
reduced, and growth retardation is rendered Avonce et al. 2004. In more general about the linking of trehalose and ABA is suggested by
Brodmann 2006. He revealed that application of 25 mM trehalosevalidamycin incresease or decrease the expression of 2277 genes, and more than a third of
them are regulated by ABA, linking trehalose to ABA or ABA signaling. Interestigly, on ABA deficient mutant abi-1 the application of trehalose and validamycin affect
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