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Plant Signal Behav
2013 Aug 01;88:. doi: 10.4161/psb.24937.
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Herbivory of maize by southern corn rootworm induces expression of the major intrinsic protein ZmNIP1;1 and leads to the discovery of a novel aquaporin ZmPIP2;8.
Lawrence SD
,
Novak NG
,
Xu H
,
Cooke JE
.
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Aquaporins channel water and other neutral molecules through cell membranes. Aquaporin gene expression is subject to transcriptional control and can be modulated by factors affecting water balance such as salt, abscisic acid and drought. During infestation of maize by southern corn rootworm (SCR), an insect that chews into and significantly damages maize roots, three maize aquaporins were differentially expressed upon prolonged infestation. Using a brief infestation of maize roots ZmNIP1;1 transcript abundance again increased under infestation while expression of a new aquaporin, ZmPIP2;8 and ZmTIP2;2 expression did not change. Since ZmPIP2;8 has not been described previously, the deduced protein sequence was analyzed in silico and found to contain the hallmarks of plant aquaporins, with a predicted protein structure similar to other functionally characterized PIP2s. NIPs characterized to date have been implicated in facilitating the movement of a variety of small molecules, while TIPs and PIPs often have the capacity to facilitate trans-membrane movement of water. Functional assays (using heterologous expression in Xenopus laevis oocytes) of ZmTIP2;2 and ZmPIP2;8 confirmed that these aquaporins demonstrate water channel capacity.
Figure 1. SCR infestation of maize roots significantly induced ZmNIP1;1 and ZmRIP2 (AAF60304) but did not affect ZmPIP2;8 and ZmTIP2;2. Plants were grown, harvested and RNA was isolated from maize roots. cDNA was synthesized and qRT-PCR performed as described in Lawrence et al.7 Relative Quantity (RQ) was calculated where control uninfested RNA at time 0 was designated as the calibrator and scaled to a value of 1. The data are expressed as log10; the log10 transformed value of the control is zero. Error bars represent standard deviation. Three biological replicates are averaged for each time point. Four plants were pooled for each biological replicate.
Figure 2. Alignment of ZmPIP2;8 and other ZmPIP2 sequences showing the conserved NPA domains of loop B in bold, loop E in bold italics and both underlined.11 The transmembrane domains are labeled TM1-6.The conserved H of loop D is starred. The conserved Ar/R filter is in red. ZmPIP2;1(AAK26758.1), ZmPIP2;2 (AAK26759.1), ZmPIP2;3 (AAK26760.1), ZmPIP2;4 (AAK26761.1), ZmPIP2;5 (AAD28761.1), ZmPIP2;6 (AAK26762.1), ZmPIP2;7 (AAK26763.1).
Figure 3. ZmPIP2;8 clusters with other ZmPIP2s in a phylogenetic tree comparing ZmPIP2;1(AAK26758.1), ZmPIP2;2 (AAK26759.1), ZmPIP2;3 (AAK26760.1), ZmPIP2;4 (AAK26761.1), ZmPIP2;5 (AAD28761.1), ZmPIP2;6 (AAK26762.1), ZmPIP2;7 (AAK26763.1),ZmPIP1;3/1;4 (Q9AQU5.1). The maximum likelihood method was used as described in the materials and methods.
Figure 4. The water permeability of the oocytes microinjected with the cRNAs of ZmPIP2;8 and ZmTIP2;2, the cRNA of Laccaria bicolor aquaporin JQ585595 as the positive control and nuclease-free water as the negative control. Swelling assay was conducted in 0.2× hypotonic MBM after 48 h incubation in MBM solution at 18 °C. Bar represents the mean of 10 replicates and error bar represents standard deviation. Letters indicate significant difference at p < 0.05.
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