Physiological Basis of Smut Infectivity in the Early Stages of Sugar Cane Colonization.

Sugar cane smut (Sporisorium scitamineum) interactions have been traditionally considered from the plant's point of view: How can resistant sugar cane plants defend themselves against smut disease? Resistant plants induce several defensive mechanisms that oppose fungal attacks. Herein, an overall view of Sporisorium scitamineum's mechanisms of infection and the defense mechanisms of plants are presented. Quorum sensing effects and a continuous reorganization of cytoskeletal components, where actin, myosin, and microtubules are required to work together, seem to be some of the keys to a successful attack.

Increase in cytotoxic lignans production after smut infection in sugar cane plants.

Smut infection alters the transcription of dirigent proteins (DIR) by sugarcane plants. Here, we show that these alterations are associated to an elevated production of cytotoxic lignans. Smut-resistant sugarcane varieties display a fivefold increase in pinoresinol and also produce elevated amounts of secoisolariciresinol. Conversely, smut-sensitive varieties do not produce pinoresinol or secoisolariciresinol upon infection, synthesizing instead small amounts of matairesinol. Our data indicate that commercial pinoresinol and secoisolariciresinol seem to prevent smut teliospore germination and sporidia release from sprouted teliospores. Consistently, we observed abundant morphological alterations of sporidia incubated in the presence of these lignans. However, commercial lignans do not block the development of the pathogen in a definitive way. Additional experiments demonstrate that only the extracts from healthy or smut-exposed resistant plants inhibit sporidia growth in vitro, indicating that a specific mixture of lignans from resistant plants is necessary to constitute an effective defense mechanism.

Polyamines levels increase in smut teliospores after contact with sugarcane glycoproteins as a plant defensive mechanism.

Previous studies have already highlighted the correlation between Sporisorium scitamineum pathogenicity and sugarcane polyamine accumulation. It was shown that high infectivity correlates with an increase in the amount of spermidine, spermine and cadaverine conjugated to phenols in the sensitive cultivars whereas resistant plants mainly produce free putrescine. However, these previous studies did not clarify the role of these polyamides in the disorders caused to the plant. Therefore, the purpose of this research is to clarify the effect of polyamines on the development of smut disease. In this paper, commercial polyamines were firstly assayed on smut teliospores germination. Secondly, effects were correlated to changes in endogenous polyamines after contact with defense sugarcane glycoproteins. Low concentrations of spermidine significantly activated teliospore germination, while putrescine had no activating effect on germination. Interestingly, it was observed that the diamine caused nuclear decondensation and breakage of the teliospore cell wall whereas the treatment of teliospores with spermidine did not induce nuclear decondensation or cell wall breakdown. Moreover, the number of polymerized microtubules increased in the presence of 7.5 mM spermidine but it decreased with putrescine which indicates that polyamines effects on Sporisorium scitamineum teliospore germination could be mediated through microtubules interaction. An increased production of polyamines in smut teliospores has been related to sugarcane resistance to the disease. Teliospores incubation with high molecular mass glycoproteins (HMMG) from the uninoculated resistant variety of sugarcane, Mayari 55-14, caused an increase of the insoluble fraction of putrescine, spermidine and spermine inside the teliospore cells. Moreover, the level of the soluble fraction of spermidine (S fraction) increased inside teliospores and the excess was released to the medium. The HMMG glycoproteins purified from Mayarí 55-14 plants previously inoculated with the pathogen significantly increased the levels of both retained and secreted soluble putrescine and spermidine. Polyamines levels did not increase in teliospores after incubation with HMMG produced by non resistant variety Barbados 42231 which could be related to the incapacity of these plants to defend themselves against smut disease. Thus, a hypothesis about the role of polyamines in sugarcane-smut interaction is explained.

Differential expression of SofDIR16 and SofCAD genes in smut resistant and susceptible sugarcane cultivars in response to Sporisorium scitamineum.

Proteomic profiling of the stalk of a smut resistant and a susceptible sugarcane cultivars revealed the presence of dirigent and dirigent-like proteins in abundance in the pool of high molecular mass (HMMG) and mid-molecular mass (MMMG) glycoproteins, produced as part of the defensive response to the fungal smut pathogen. Quantitative RT-PCR analysis showed that expression levels of SofDIR16 (sugarcane dirigent16) and SofCAD (sugarcane cinnamyl alcohol dehydrogenase) were higher in the smut resistant My 55-14 cultivar than in the sensitive B 42231 cultivar prior to infection. Inoculation with fungal sporidia or water decreased the level of SofCAD transcripts in My 55-14, indicating that regulation of SofCAD expression does not take part of the specific response to smut infection. In contrast, SofDIR16 expression was almost nullified in My 55-14 after inoculation with fungal sporidia, but not after water injection. It is proposed that the decreased expression of dirigent proteins induces the formation of lignans, which are involved in the defense response of the smut resistant My 55-14 cultivar.

Defence sugarcane glycoproteins disorganize microtubules and prevent nuclear polarization and germination of Sporisorium scitamineum teliospores.

Microtubules (MTs) are involved in the germination of Sporisorium scitamineum teliospores. Resistant varieties of sugar cane plants produce defence glycoproteins that prevent the infection of the plants by the filamentous fungi Sporisorium scitamineum. Here, we show that a fraction of these glycoproteins prevents the correct arrangement of MTs and causes nuclear fragmentation defects. As a result, nuclei cannot correctly migrate through the growing hyphae, causing germinative failure. Arginase activity contained in defence glycoproteins is already described for preventing fungal germination. Now, its enzymatically active form is presented as a link between the defensive capacity of glycoproteins and the MT disorganization in fungal cells. Active arginase is produced in healthy and resistant plants; conversely, it is not detected in the juice from susceptible varieties, which explains why MT depolarization, nuclear disorganization as well as germination of teliospores are not significantly affected by glycoproteins from non-resistant plants. Our results also suggest that susceptible plants try to increase their levels of arginase after detecting the presence of the pathogen. However, this signal comes "too late" and such defensive mechanism fails.