Alice Cheung and Hen-ming Wu's laboratory

Our overall interest is on plant cell signal transduction. We have two focus areas.

1. The functional role of formins, proteins that nucleate actin polymerization, in regulating plant actin cytoskeleton, growth and development

Plant actin cytoskeleton supports multiple cellular processes that underlie growth and development. For example, the establishment of auxin gradients, which is fundamental to plant growth and differentiation, cell plate formation during cell division, tip growth in root hairs and pollen tubes, intracellular organelle movements, such as trafficking of Golgi bodies and transport vesicles, and the movement of chloroplasts in response to light, are all dependent on the actin cytoskeleton. Therefore, the actin cytoskeleton impacts developmental processes from embryogenesis to fertilization, and how plants respond to the environment. Actin-nucleation, the polymerization of the first two to three monomeric actins, is the rate-limiting step in the formation of actin filaments, structures that underlie the actin cytoskeleton. Formins are one of two known families of actin-nucleating proteins. One of the research focus in our lab is to determine the functional roles formins in plant growth and development. Some plant formins (group I) have the plant-unique feature of having a potentially glycosylated extracellular and a transmembrane domain that anchor the cytosolic actin-interacting domain to the cell surface. Group I formins thus serve as perfect candidates for mediating extracellular stimuli directly to the actin cytoskeleton, analogous to integrins in other organisms but which have not been identified in plants. We are using molecular, cell biological, and reverse genetics approaches to elucidate the functional roles for AFHs in plant growth and development. These studies will provide linkages between a critical biochemical activity that regulates actin polymerization to growth and development. We are also identifying interacting partners for formins so as to understand how these proteins are regulated.

For references for this and related projects:

1. Cheung, A.Y., and Wu, H-M. (2004) Over-expression of an Arabidopsis Formin stimulates supernumerary actin cable formation from pollen tube cell membrane. Plant Cell 16: 257-269.

2. Cheung, A.Y., Chen, C., Glaven, R., Vidali, L., Hepler, P.K., and Wu, H-M. (2002) Rab2 regulate vesicular transport between endoplasmic reticulum and Golgi bodies and is important for pollen tube elongation. Plant Cell, 14: 945-962.

3. Chen, C., Wong, E., Vidali, L., Estavillo, A., Hepler, P.K., Wu, H-M., and Cheung, A.Y. (2002) The regulation of actin organization by actin depolymerizing factor (ADF) in elongating pollen tubes. Plant Cell 14: 2175-2190.

4. Chen, C. Y-h., Cheung, A.Y. and Wu, H-M. (2003). Actin depolymerizing factor mediates Rac/Rop GTPase regulated pollen tubegrowth. Plant Cell 15: 237-249.

5. De Graaf, B., Cheung, A.Y., Andreyeva, T., Levasseur, K., Kiliesheski, M., Wu, H-M., (2005) Rab11 GTPase-regulated membrane trafficking is crucial for tip-focused pollen tube growth. Plant Cell (almost in press, pending minor revisions 5-05).

2. Small GTPases in plant signaling pathways

Rac and Rab GTPases are molecular switches that regulate diverse cellular processes. We are interested in understanding their functional roles in hormone signaling. The auxin indole-3-acetic acid (IAA) is a key plant hormone essential for a broad range of growth and developmental processes. Auxin-signaled activation of responsive genes by 26S proteasome-dependent proteolysis of a family of short-lived repressor proteins, Aux/IAA, is a most extensively examined system of regulation via regulated proteolysis. However, mechanisms pertaining to reception and transduction of the auxin signal to intracellular signaling pathways remain unclear. We have observed that a subset of Rac GTPases is activated by auxin and they mediate the auxin signal to activate downstream responsive genes in transfected protoplasts and in whole plants. Moreover, Rac GTPases also increase the instability in Aux/IAA proteins whereas overproduction of Aux/IAA proteins suppresses Rac-induced auxin-responsive gene expression. These observations suggest that RacGTPases play an integral role in the auxin-induced signaling pathway of gene derepression via proteolysis. Even more exciting is our discovery that auxin and the substrate Aux/IAA proteins induce recruitment of proteolytically active nuclear protein particles where degradation of these repressors occur and this process is also mediated by Rac GTPases.

Multiple Arabidopsis AtRACs have the redundant ability to mediate the auxin signal to downstream responsive genes. Together they are likely to participate in regulating auxin response throughout plant development. One of our current research areas is to determine how individual AtRACs contribute to auxin signaling on the tissue, organ and developmental levels. We are using molecular, cell biological and reverse genetics approaches to understand how these highly homologous proteins mediate opposite functions. Attempts will be made to identify their interacting molecules towards understand how they mediate diverged functions.

For references on this work:

1. Tao, L-z., Cheung, A.Y., and Wu, H-M. (2002) Plant Rac-like GTPases are activated by auxin and mediate auxin responsive gene expression. Plant Cell 14: 2745-2760.
2. Tao, L-z., Cheung, A.Y., Nibau, C., and Wu, H-M. (2005) Rac GTPases mediate auxin-induced formation of Aux/IAA-containing proteolytically active nuclear protein bodies. Plant Cell, 17: 2369-2383.

• Nibau, C., Wu, H-M. and Cheung, A.Y. (2006) RAC/ROP GTPases: “Hubs” for signal integration and diversification in plants. Trends in Plant Science, 11, 1360-1385.

Our Rab GTPase project is not described here, but it is becoming evident that vesicle trafficking, which Rab GTPases regulate, are intimately related to many intracellular signaling pathways. Part of our efforts is to connect the pathways regulated by these two classes of small GTPases and how they together regulate the actin cytoskeleton.

For references on Rab GTPase-related work:

1. Cheung, A.Y., Chen, C., Glaven, R., Vidali, L., Hepler, P.K., and Wu, H-M. (2002) Rab2 regulate vesicular transport between endoplasmic reticulum and Golgi bodies and is important for pollen tube elongation. Plant Cell, 14: 945-962.

2. de Graaf, B. H.J., Cheung, A.Y., Andreyeva, T, Levasseur, K., Kieliszewski, M., and Wu, H-M. (2005) Rab11 GTPase-Regulated Membrane Trafficking Is Crucial for Tip-Focused Pollen Tube Growth in Tobacco. Plant Cell 17: 2564-2579.