Identification and functional characterization of the Arabidopsis Snf1‐related protein kinase SnRK2.4 phosphatidic acid‐binding domain

Phosphatidic acid (PA) is an important signalling lipid involved in various stress‐induced signalling cascades. Two SnRK2 protein kinases (SnRK2.4 and SnRK2.10), previously identified as PA‐binding proteins, are shown here to prefer binding to PA over other anionic phospholipids and to associate with cellular membranes in response to salt stress in Arabidopsis roots. A 42 amino acid sequence was identified as the primary PA‐binding domain (PABD) of SnRK2.4. Unlike the full‐length SnRK2.4, neither the PABD‐YFP fusion protein nor the SnRK2.10 re‐localized into punctate structures upon salt stress treatment, showing that additional domains of the SnRK2.4 protein are required for its re‐localization during salt stress. Within the PABD, five basic amino acids, conserved in class 1 SnRK2s, were found to be necessary for PA binding. Remarkably, plants overexpressing the PABD, but not a non‐PA‐binding mutant version, showed a severe reduction in root growth. Together, this study biochemically characterizes the PA–SnRK2.4 interaction and shows that functionality of the SnRK2.4 PABD affects root development.     

Differential responses of mosquito sibling species Anopheles arabiensis and An. quadriannulatusto carbon dioxide, a man or a calf

Field studies on responses of two mosquito sibling species, Anopheles arabiensis Patton and An. quadriannulatus Theobald, to a man, a calf and different release rates of carbon dioxide (man, calf and cow equivalents) were conducted in north-eastern South Africa. Various combinations of baits were compared in two-choice tests, using two mosquito nets, placed 2.5 m apart and 10 cm off the ground. Mosquitoes attracted to the baits were able to enter the nets from below and were collected by means of a suction tube. In a two-choice test between a man and CO₂ (human equivalent, 250 ml/min), 81% of the An. quadriannulatus were caught with CO₂. The reverse was seen for An. arabiensis , where only 20% of the total catch was caught with CO₂ compared to man. High release rates of CO₂ (cow equivalent, 800 ml/min) attracted significantly more An. quadriannulatus than the low release rate (250 ml/min), whereas no significant effect of the release rate of CO₂ on the total catch of An. arabiensis was seen. In the latter species, up to 33% of the attraction of human emanation is attributable to carbon dioxide. Anopheles quadriannulatus was equally attracted to a calf and CO₂ (calf equivalent, 180 ml/min). Catches of other mosquito species showed consistent differences between all treatments which appear to be associated with differences in host-preference, suggesting that the importance of CO₂ in host-seeking behaviour of mosquitoes increases with the degree of zoophily.     

Reassessing the role of phospholipase D in the Arabidopsis wounding response

Plants respond to wounding by means of a multitude of reactions, with the purpose of stifling herbivore assault. Phospholipase D (PLD) has previously been implicated in the wounding response. Arabidopsis ( Arabidopsis thaliana ) AtPLD α 1 has been proposed to be activated in intact cells, and the phosphatidic acid (PA) it produces to serve as a precursor for jasmonic acid (JA) synthesis and to be required for wounding-induced gene expression. Independently, PLD activity has been reported to have a bearing on wounding-induced MAPK activation. However, which PLD isoforms are activated, where this activity takes place (in the wounded or non-wounded cells) and what exactly the consequences are is a question that has not been comprehensively addressed. Here, we show that PLD activity during the wounding response is restricted to the ruptured cells using ³²P_i-labelled phospholipid analyses of Arabidopsis pld knock-out mutants and PLD -silenced tomato cell-suspension cultures. pldα1 knock-out lines have reduced wounding-induced PA production, and the remainder is completely eliminated in a pldα1 / δ double knock-out line. Surprisingly, wounding-induced protein kinase activation, AtLOX2 gene expression and JA biosynthesis were not affected in these knock-out lines. Moreover, larvae of the Cabbage White butterfly ( Pieris rapae ) grew equally well on wild-type and the pld knock-out mutants.