Our interest in HDACs derives from the discovery that HC-toxin, a critical virulence determinant for the plant pathogenic fungus Cochliobolus carbonum, is a potent and specific inhibitor of HDACs from maize and other organisms. This has led us to study HDAC function in filamentous fungi and plants, especially in relation to pathogenesis. This work has been a collaboration with Peter Loidl, Gerald Brosch, and Stefan Graessle at the University of Innsbruck, Austria.
Four Research Questions:
Why does inhibition of maize HDACs permit the development of disease?
C. carbonum is an exceptionally virulent pathogen on susceptible maize (Figure 1). The cyclic tetrapeptide HC-toxin plays the central role in this disease: Maize that is insensitive to HC-toxin is resistant to C. carbonum, and isolates of C. carbonum that do not makeHC-toxin are avirulent. Unlike other host-selective toxins, however, HC-toxin is not toxic to plant (or animal) cells (Wolf and Earle, 1991). Thus, simple killing of its host cannot explain the exceptional virulence of C. carbonum.
HC-toxin is a specific inhibitor of histone deacetylases of the RPD3/HDA1 classes (Brosch et al., 1995). This class of HDAC is found in all eukaryotes including yeast, mammals, and plants. Clearly, HDACs have a a critical role in defense of plants (at least maize) against pathogens. One hypothesis is that HC-toxinis a suppressor of HDAC-regulated defense responses (Brosch et al., 1995), but, if so, the nature of those responses is not known.
Understanding the role of HC-toxin in the disease process depends on our understanding of HDAC function in general. This has proven to be a 'moving target'. First, plants, like other organisms, have multiple HDACs of the RPD3/HDA1 class (at least 15 each in Arabidopsis and maize), and all are apparently sensitive to HC-toxin. Second, HDACs have many substrates. In addition to histones, HDACs can deacetylate other proteins including a number of transcription factors such as NF-kappaB (Chen et al., 2001). Tubulin is also a substrate for a specific human HDAC.
What virulence functions in C. carbonum are regulated by HDACs?
One of the HDACs of C. carbonum, HDC1, is required for full virulence. An hdc1 mutant shows reduced growth on alternate carbon sources, reduced expression of cell wall degrading enzymes, and reduced leaf penetration efficiency (Baidyaroy et al., 2001) (Figure 2). What are the critical virulence factors controlled by HDC1- are they cell wall degrading enzymes, either individually or collectively, or perhaps unrelated biochemical processes?
How does C. carbonum protect itself against its own potent HDAC inhibitor?
Having HDACs, the fungus must have a mechanism to keep from killing itself when making HC-toxin. We have discovered that whereas the HDAC activity of most other organisms,including Aspergillus nidulans andNeurospora crassa, are fully sensitive to HC-toxin and trichostatin, the HDAC activities of some C. carbonumisolates are resistant.
Anion exchange fractionation of the HDAC activity of filamentous fungi reveals two major peaks of activity, one that is resistant and one that is sensitive to HC-toxin (Brosch et al., 2001). There is evidence for both intrinsic and extrinsic resistance mechanisms. For example, extracts from resistant C. carbonum can protect HDAC activity in extracts from sensitive strains (Baidyaroy et al., 2002).
- Is HDAC inhibition by plant pathogenic fungi a general virulence strategy?
A number of fungi make HDAC inhibitors. Several of them are, or are closely related to, plant pathogens (Figure 3). The existence of these HDAC inhibitors raises the intriguing possibility that HDAC inhibition is a virulence strategy not restricted to C. carbonum. The gene cluster for apicidin (Fig. 3) was recently described from Fusarium semitectum. Its genes are highly homologous to those of TOX2 controlling HC-toxin biosynthesis (Jin et al., Mol Microbiol 76:456-466).
References on HDACs and HC-toxin:
Baidyaroy, D., G. Brosch, J.-H. Ahn, S.Grassle, S. Wegener, O. Caballero, P. Loidl, and J.D. Walton (2001) A gene related to yeast HOS2 is necessary for extracellular depolymerase expression and virulence in a plant pathogenic fungus. Plant Cell 13:1609-1624.
Baidyaroy, D., G. Brosch, S. Graessle, P. Trojer, and J.D. Walton (2002) Characterization of inhibitor-resistant histone deacetylase activity in plant-pathogenic fungi. Eukaryotic Cell 1:538-547.
Brosch, G., R. Ransom, T. Lechner, J.D. Walton, and P. Loidl (1995) Inhibition of maize histone deacetylase by HC-toxin, the host-selective toxin of Cochliobolus carbonum. Plant Cell 7:1941-1950.
Brosch, G., M. Dangl, S. Graesle, A. Loidl, P. Trojer, E.-M. Brandtner, K. Mair, J.D Walton, D. Baidyaroy, and P. Loidl (2001) An inhibitor-resistant histone deacetylase in the plant pathogenic fungus Cochliobolus carbonum.Biochemistry 40:12855-12863.
Graessle, S., M. Dangl, H. Haas, K. Mair, P. Trojer, E.-M.Brandtner,J.D. Walton, P. Loidl, and G. Brosch (2000) Characterization of two putative histone deacetylase genes from Aspergillus nidulans. Biochim. Biophys. Acta 93405:1-7.
Lechner, T., A. Lusser, A. Pipal, G. Brosch, A. Loidl, M. Goralik-Schramel,R. Sendra, S. Wegener, J. D. Walton, and P. Loidl (2000) RPD3-type histone deacetylases in maize embryos. Biochemistry 39:1683-1692.
Ransom, R.F., and J.D. Walton. (1997) Histone hyperacetylation in maize in response totreatment with HC-toxin or infection by Cochliobolus carbonum. Plant Physiol. 115:1021-1027.
Wolf S.J., and Earle, E.D. (1991). Effects of Helminthosporium carbonum race 1 toxin on host and non-host cereal protoplasts. Plant Sci. 70: 127-137.
Some HDAC papers from other labs:
Kijima, M., Yoshida, M., Suita, K., Horinouchi, S., and Beppu, T. (1993) Trapoxin, an antitumor cyclic tetrapeptide, is an irreversible inhibitor of mammalian histone deacetylase. J. Biol. Chem. 268, 22429-22435.
Kwon, H.J., Owa, T., Hassig, C.A., Shimada, J., and Schreiber, S.L.(1998) Depudecin induces morphological reversion of transformed fibroblasts via the inhibition of histone deacetylase. Proc. Natl. Acad. Sci. U.S.A. 95, 3356-3361
Selker, E.U. (1998) Trichostatin A causes selective loss of DNA methylation in Neurospora. Proc. Natl. Acad. Sci. U.S.A. 95, 9430-9435.
Bernstein, B.E., Tong, J.K., and Schreiber, S.L. (2000) Genomewidestudies of histone
deacetylase function in yeast. Proc. Natl. Acad. Sci. U.S.A.97, 13708-13713.
Jenuwein, T., and C.D. Allis (2001) Translating the histone code. Science 293:1074-1080.
Lo, W.-S., L. Duggan, N.C. Tolga Emre, R. Belotserkovskya,W.S. Lane, R. Shiekhattar, and S.L. Berger (2001) Snf1 - a histone kinase that works in concert with the histone acetyltransferase Gcn5 to regulate transcription.Science 293:1142-1146.