CBP60b clade proteins are prototypical transcription factors mediating immunity.

Transcriptional reprogramming is critical for plant immunity. Several calmodulin (CaM)-binding protein 60 (CBP60) family transcription factors (TFs) in Arabidopsis (Arabidopsis thaliana), including CBP60g, Systemic Acquired Resistance Deficient 1 (SARD1), CBP60a, and CBP60b, are critical for and show distinct roles in immunity. However, there are additional CBP60 members whose function is unclear. We report here that Arabidopsis CBP60c-f, four uncharacterized CBP60 members, play redundant roles with CBP60b in the transcriptional regulation of immunity responses, whose pCBP60b-driven expression compensates the loss of CBP60b. By contrast, neither CBP60g nor SARD1 is inter-changeable with CBP60b, suggesting clade-specific functionalization. We further show that function of CBP60b clade TFs relies on DNA-binding domains (DBDs) and CaM-binding domains, suggesting that they are downstream components of calcium signaling. Importantly, we demonstrate that CBP60s encoded in earliest land plant lineage Physcomitrium patens and Selaginella moellendorffii, are functionally homologous to Arabidopsis CBP60b, suggesting that the CBP60b clade contains the prototype TFs of the CBP60 family. Furthermore, tomato and cucumber CBP60b-like genes rescue the defects of Arabidopsis cbp60b and activate the expression of tomato and cucumber SALICYLIC ACID INDUCTION DEFICIIENT2 (SID2) and ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) genes, suggesting that immune response pathways centered on CBP60b are also evolutionarily conserved. Together, these findings suggest CBP60b clade transcription factors are functionally conserved in evolution and positively mediate immunity.

Canonical Rab5 GTPases are essential for pollen tube growth through style in Arabidopsis.

Pollen tubes have dynamic tubular vacuoles. Functional loss of AP-3, a regulator of one vacuolar trafficking route, reduces pollen tube growth. However, the role of canonical Rab5 GTPases that are responsible for two other vacuolar trafficking routes in Arabidopsis pollen tubes is obscure. By using genomic editing, confocal microscopy, pollen tube growth assays, and transmission electron microscopy, we demonstrate that functional loss of canonical Rab5s in Arabidopsis, RHA1 and ARA7, causes the failure of pollen tubes to grow through style and thus impairs male transmission. Functional loss of canonical Rab5s compromises vacuolar trafficking of tonoplast proteins, vacuolar biogenesis, and turgor regulation. However, rha1;ara7 pollen tubes are comparable to those of wild-type in growing through narrow passages by microfluidic assays. We demonstrate that functional loss of canonical Rab5s compromises endocytic and secretory trafficking at the plasma membrane (PM), whereas the targeting of PM-associated ATPases is largely unaffected. Despite that, rha1;ara7 pollen tubes contain a reduced cytosolic pH and disrupted actin microfilaments, correlating with the mis-targeting of vacuolar ATPases (VHA). These results imply a key role of vacuoles in maintaining cytoplasmic proton homeostasis and in pollen tube penetrative growth through style.

Vesicle trafficking in Arabidopsis pollen tubes.

The delivery of sperm cells via tip-growing pollen tubes is an innovation of seed plants and shows the importance of pollen tubes for reproduction and their specific growth kinetics. Fast-growing pollen tubes demand an extensive and dynamic vesicular trafficking network to build new cell membrane and wall, to deliver proteins among endomembrane compartments, and also to respond to external stimuli for growth adjustment. In this review, we summarize current findings on endomembrane compartments and vesicular trafficking routes of pollen tubes, comparing and contrasting their features with those of most somatic cells. We discuss the importance of membrane homeostasis, either at the plasma membrane (PM) or between PM-targeted trafficking and vacuolar trafficking, for pollen tube growth. We also provide perspectives to facilitate future studies of vesicular trafficking in pollen tubes.