Processes Underlying Fruit Cuticle Integrity in Tomato (Solanum spp.)
The ability to synthesize, deposit and maintain a biomechanically resilient epidermis and a hydrophobic cuticle covering aerial organs are crucial protective features of all land plants. To achieve an intact and dynamic epidermal layer in growing tissues, the deposition of complex primary cell wall polysaccharides and structural cuticular lipids must involve complex and coordinated trafficking, assembly, and restructuring processes. In rapidly expanding organs, such as fleshy fruits, lack of coordination of these processes can result in cuticle and epidermal defects, manifested as cracking and corky scarring, which in crops can lead to substantial economic losses. Identification of the causes of such defects can also provide new insights into the molecular basis of systems to maintain epidermal integrity. As an example, the tomato HYPERCRACKING 1 (hcr1) mutant shows extensive fruit cracking at an early developmental stage, and consequent massive deposition of suberin in the epidermis during growth. Map-based cloning of the hcr1 locus and biochemical analysis suggested that the hyper-cracking phenotype is a consequence of defective sterol biosynthesis. Morphological, cytological, and molecular characterization of hcr1 fruit revealed inhibited cell expansion and division in the pericarp and reduced cellulose levels leading to impaired pericarp development, resulting in uncoordinated expansion between the pericarp and the inner tissues leading to cracking. This study suggests that sterols are important for primary cell wall deposition, and notably cellulose synthesis. Moreover, the presence of a strong cracking phenotype in fruit, but not vegetative tissues, highlights variability in the susceptibility of different organs to impaired epidermal integrity.Relatedly, a second study is concerned with microscopic pores associated with trichomes on the fruit cuticle that are exposed when trichomes are dislodged. Staining of the pore areas with Toluidine Blue has shown that they have the capacity to become sealed, an adaptation with implications for water retention and relevant to open research questions regarding cuticle structure and remodeling. The South American tomato relative Solanum quitoense (common name: naranijlla) displays large, abundant, and uniformly spaced trichomes on the fruit surface, and here we used this expedient model for studying the sealing process. Evapotranspiration experiments revealed that removal of trichomes at harvest significantly increases fruit water loss, confirming that the trichome pores are significant routes of water loss, as in S. lycopersicum. However, isolated cuticles that have been allowed to seal prior to isolation allow less transcuticular water loss than those isolated immediately after trichome removal, indicating that pore sealing effectively inhibits this water loss through pores. Staining the cuticle with Toluidine Blue and imaging the surface revealed that the area of the pore stain gets measurably smaller with sealing time, plateauing at approximately 3 days. Viewing these stained pores at cross section, contextualized to SEM images, suggests that the sealing does not occur at the surface of the trichome scar, but rather along the base of the exposed cavity. Evidence of the sealing mechanism was not captured by confocal microscopy and staining with Basic Fuchsin, Fluorol Yellow, and Calcofluor White, possibly due to the complexity of the S. quitoense cuticle architecture and the lack of specificity of stains primarily characterized in Arabidopsis. This rules out, however, that the sealing mechanism involves a large (>10 micrometer) material deposit, as this would have been detectable. Additional study will be necessary to understand the mechanism of this sealing process and to determine whether these findings generalize to tomatoes of greater commercial interest, especially S. lycopersicum.