UC Riverside

Stem-cell homeostasis is mediated by multifaceted networks involving plant hormones and local cell-cell communication. Earlier studies have implicated cytokinins in regulating shoot apical meristem (SAM) growth. However, the precise role of cytokinin in SAM remains largely unknown because cytokinins have been implicated in several developmental processes and merely studying the terminal phenotypes may not reveal their function in actively developing SAMs. The live-image technology allows studying the function of any given regulator immediately after its perturbations in transient experiments. The live-imaging work presented here provides: 1) Functional analysis of SAM-enriched cytokinin activating enzymes, 2) The live-image study on key regulators in cytokinin biosynthesis and signaling, 3) The function of SHOOTMERISTEMLESS (STM).

The cell-type specific genomics has predicted and RNA in situ analysis has confirmed the enrichment of cytokinin activating enzymes in the SAM-stem cell niche. Knock-out three centrally expressed members alters phyllotaxy. Transient manipulation of cytokinin results in organ positioning defects within the peripheral zone (PZ) of SAMs. The levels of cytokinin are correlated to auxin responses. Moreover, external application of auxin fails to induce auxin responses suggesting that cytokinins are required for activating auxin response pathway components. Cell division analyses reveal that cell mitotic activities are correlated to the levels of cytokinins. We conclude that cytokinin controls phyllotaxy by regulating auxin signaling, auxin transport and cell division rates.

We find that WUSCHEL (WUS) regulates a type-B response regulator, ARR1. Then, we have studied the effects of positive cytokinin signaling by using inducible activation of constitutively active forms of ARR1 coupled with live-image technology. This analysis has revealed that constitutive activation of cytokinin signaling results in expansion of stem-cell domain leading to de-differentiation of differentiating cells and this process is WUS dependent.

Genetically, stm mutant phenotype implies that STM function is required for SAM initiation and fulfills a complementary role to that of WUS. Inducible inactivation of STM has shown that it is required for preventing the CZ cells from responding to auxin. Furthermore, other differentiation markers were also found to be mis-expressed in the CZ cells. These findings lead to conclude that STM prevent the stem-cell domain from differentiation.