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CELL WALL-ASSOCIATED KINASES Cell wall associated Kinases (WAKs) family is one of the candidate for a cell-wall–cytoplasmic signaling molecules. The family of Arabidopsis cell wall associated serine–threonine kinases (WAKs) are linked directly to the cell wall, and they span the plasma membrane and have a cytoplasmic kinase domain [1]. The cell wall-associated kinases (WAKs) are receptor-like kinases that are attached to pectin component of the cell wall, and have a cytoplasmic protein kinase domain. WAKs’ association with cell wall is very strong (having covalent link to pectin), such that its release from the cell wall requires enzymatic digestion [1]. Under conditions that collapse the turgor of a plant cell so as to separate the membrane from the wall (plasmolysis), the WAKs-wall association is so strong that they remain in the cell wall. There are five WAK’s isoform in Arabidosis with variable extracellular domain within these isoform [1], all of which contain at least two epidermal growth factor (EGF). Of all these isoforms, WAK1 and WAK2 are most ubiquitous and their messenger RNA (mRNAs) and proteins are present in vegetative meristem and areas of cell expansion [1].

WAKs’ Identification.

Boiling of cells in detergent has been reported to be the only means of extracting WAKs cross-linkage in insoluble materials. All the five WAKs’ genes are tightly clustered in tandem in a 30-kb locus on chromosome1 [2]. Furthermore, the chance isolation of WAKs’ cDNAs is the means through which the WAKs were identified, and such identification subsequently unveils their significance to the cell wall. In addition, electron micrographs with specific WAK-kinase antiserum has been reported to show WAK protein in the cell wall and on the plasma membrane. Yet protease treatment of protoplasts and Western blots revealed that the kinase domain was cytoplasmic, and the receptor crossed the membrane such that the epidermal growth factor (EGF) domain is placed in the extracellular space [1, 2]. However, their encoded kinase domains have been shown to be 85% identical, and their extracellular regions display up to 65% identity [1, 2]. Notably, all WAKs contain the same conserved spacing of cysteine residues in the extracellular domain, being implicated as the hallmark of the EGF repeat. [4].

WAKS are required for cell expansion. The WAK genes have been severally reported to play vital role in cell expansion/cell elongation [5, 6]. The WAK 2 null allele have been shown to cause a loss of expansion in root, particularly under limited sugar and salt conditions [7]. The reduction in vacuolar invertase activity at the root of WAK2-1 null mutant, suggested that WAKs regulates invertase at transcriptional level, and these results support a model where WAKs regulate cell expansion by control of sugar concentration and probably tugor control [8].

WAKs binds and are responsive to pectin.

As shown by electron microscopy, WAKs lies in the cell wall (cross-linked to cell wall materials). The pectinases release WAKs from cell wall materials, this leads to initial suggestion that WAKs are bound to Pectins (Anderson et al, 2001, Wagner and Kohom, 2001) [6,9]. Notably, the extracted WAK Protein is still bound to pectin epitope on denaturing gels, this suggests its covalent binding to a peptic fragment [9]. In the same trend, the purified extracellular domains of WAK 1 and WAk2 bind to pectin in vitro, several lines of evidences have shown that the binding of pectin to WAKs activates many signaling pathways. Also, it has been reported that pectin treatment of protoplast causes the induction and repression of hundreds of genes that are involved in the cell biogenesis and stress responses, and the same response is blocked in the cells that lack WAK2 [7,10]. Besides pectin, WAKs may also bind to glycine-rich protein (GRPs) of the cell walls. The WAK-GRP3 interaction is specific to WAK1. More so, both GRP3 and WAK1 can be isolated from the same 450 KDa complex in Arabidosis extracts. But more surprisingly, WAK1 is normally not detected as water soluble protein [11].

WAKs as a cell wall sensors.

WAKs are like many other plant receptors in that they regulate some aspect of cell division or growth, but also affect aspects of cell wall; perhaps WAKs are also cell wall sensors. WAKs are also involved in pathogen and stress responses, but are likely activated in this case by fragmented pectin [1].

WAKS's interaction with oligogalacturonic acids initiates stress response.

WAKs are also involved in the response to pathogen and stress. WAKs gene expression is induced by wounding, pathogen infection, and by many stress factors such as Ozone and heavy metals [9, 12]. The oligogalacturonic acids, OGs binding to WAKs usually initiate its response to pathogen and wounding [7,10,13].

WAKs distinguish the state of pectin: a model.

In native cell walls, the Wall-associated kinases had been shown to bound to pectin, and their activity is required for normal cell expansion, [7,10,14] In summary, the results are consistent with a model described elsewhere, that the type and concentration of pectin present in the wall could lead to a WAK-dependent activation of different signaling pathways., it has been shown that the unchallenged but expanding walls would preferentially activate via WAKs, a cell expansion path that includes, Map Kinase 3, MPK3 (and that other receptors are signaling via MPK3 and 6 is indicated by additional arrows). However, when OGs are generated by a wall disturbance, the WAKs may alter their signaling path to help initiate the stress response by now also activating Map Kinase 6, MPK6 and a new downstream response [14]. The result from in vitro binding assays reveal a higher binding affinity of OGs than longer polymers for WAKs [14], this suggests a mechanism by which WAKs can switch from binding the native cell wall pectin to OGs, thereby distinguishing types of pectin. Differential activation by various pectins might be achieved by the specific pectin affinity of an individual receptor, or probably the combinations of WAKs with as yet unidentified partners [1].

In conclusion, it is well understood beyond any doubt that the plant cell wall is an extracellular matrix, ECM, and that it does indeed determines cell shape and size. More so, it also constitutes molecules that suggest that protein and carbohydrate ligands, as well as membrane receptors play a prominent role in modeling plant development. Notably, WAKs are somehow involved in signaling from the pectin extracellular matrix in coordination with GRPs, and our full knowledge of this mechanism may be of tremendous help to our understanding of the cell wall's role in cell expansion during development. It is tempting to draw a parallel between WAKs, GRPS and pectins with metazoan receptor kinases that are activated by peptide growth hormone–carbohydrate complexes [15].

References.

1. Kohorn and Susan L. Kohorn (2012). The cell wall-associated kinases, WAKs, aspect in receptors. doi: 10.3389/fpls.2012.00088.

2. He ZH, Cheeseman I, He D, Kohorn BD: A cluster of five cell wall associated receptor kinase genes, Wak1-5, are expressed in specific organs of Arabidopsis. Plant Mol Biol 1999, 39:1189-1196.

3. Z.H He, M Fujiki, B.D KohornA cell wall-associated, receptor-like kinase. J Biol Chem, 271 (1996), pp. 19789-19793.

4. Sampoli Benitez, B. A., and Komives, E. A. (2000). Disulfide bond plasticity in epidermal growth factor. Proteins 40, 168–174.

5. Lally,D.,Ingmire,P, Tong,H.Y, and He,Z.H.(2001). Antisense expression of a cell wall- associated protein kinase, WAK4, inhibits cell elongation and alters morphology. Plant Cell 13, 1317–1331.

6. T.A Wagner, B Kohorn Wall associated kinases, WAKs, are expressed throughout plant development and are required for cell expansion. Plant Cell, 13 (2001), pp. 303-318.

7. Kohorn,B.D.,Kobayashi,M.,Johansen, S., Riese,J.,Huang,L.F.,Koch,K.,Fu, S., Dotson,A.,and   Byers,N.(2006b). An Arabidopsis cell wall-associated kinase required for  and cell growth. PlantJ. 46, 307–316.

8. Kohorn BD 2006: Plasma membrane-mell wall contacts. Plant Physiol 2000, 124:31-35.

9. Anderson,C.M., Wagner,T.A.,Perret, M., He,Z.H.,He,D.,and Kohorn,B. D.(2001).WAKs:cell wall-associated kinases linking the cytoplasm to the extracellular matrix. PlantMol. Biol. 47, 197–206.

10. Kohorn, B.D.,Johansen,S.,Shishido, A., Todorova,T.,Martinez,R.,Defeo, E., and. Obregon, P.(2009).Pectin activation of MAP kinase and gene expressionisWAK2dependent Plant J. 60, 974–982.

11. Park, A.R., Cho, S.K., Yun, U.J., Jin,M. Y.,Lee,S.H.,Sachetto-Martins, G., and Park,O.K.(2001).Interaction of the Arabidopsis receptor protein kinase Wak1 with a glycine-rich pro- tein,AtGRP-3. J. Biol. Chem. 276, 26688–26693.

12. He, Z. H., He, D., and Kohorn, B. D. (1998). Requirement for the induced expression of a cell wall associated receptor kinase for survival during the pathogen response. Plant J. 14, 55–63.

13. Denoux, C.,Galletti,R.,Mammarella, N., Gopalan,S.,Werck,D.,De Lorenzo,G.,Ferrari,S., Ausubel, F.M.,and Dewdney,J.(2008).Activation of defense response pathways by OGs and Flg22 elicitors in Arabidopsis seedlings. Mol.Plant 1, 423–445.

14. Brutus,A.,Sicilia,F.,Macone,A.,Cer- vone,F.,and DeLorenzo,G.(2010). A domains wap approach reveals a role of the plant wall-associated kinase 1(WAK1) as a receptor of oligo galacturonides. Proc.Natl.

15. Buce D. Kohorn (2001). WAKs; cell wall associated kinases. Commentary. Current Opinion in cell Biology 2001, 13:529-533.