Aquaporin activity of barley tonoplast intrinsic proteins is involved in the delay of the coalescence of protein storage vacuoles in aleurone cells

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The coalescence of protein storage vacuoles (PSVs) is one of the most prominent cellular changes occurring in cereal aleurone cells during germination. This structural change is highly coupled with the functional transition of this organelle from a storage compartment to a lytic section. Gibberellic acid (GA) promotes this process, whereas abscisic acid (ABA) prevents it. Previously, we demonstrated that ABA-inducible HvTIP3;1 plays a decisive role in ABA-mediated prevention of PSV fusion. In this follow-up study, we examined whether the aquaporin activity of tonoplast intrinsic protein (TIP) is related to its preventive effect on PSV fusion using various functional mutants. The defective forms of aquaporin (HvTIP3;1m and HvTIP3;1ΔNPA-GFPs for HvTIP3;1, and HvTIP1;2m for HvTIP1;2) were found to be less effective than the usual form in delaying the PSV fusion process occurring in GA-treated cells. In contrast, overexpression of HvTIP3;1m reduced the preventive effect of ABA on PSV fusion. Upon inhibition of aquaporin activity using mercury, PSV fusion occurred to a greater extent in ABA-treated barley protoplasts. These data suggest that the aquaporin activity of TIP is involved in the deterrent effect of TIP on PSV coalescence. TIP3-GFP barley transgenic seeds showed prolonged expression of the TIP3;1 transcript. Moreover, PSV fusion progressed at a much slower rate compared to wild type. Additionally, the degradation of storage proteins was not as efficient, suggesting that a metamorphic transition of PSVs to lytic organelles is closely correlated with the disappearance of HvTIPs and the PSV fusion process.