Failure of the Tomato Trans-Acting Short Interfering RNA Program to Regulate AUXIN RESPONSE FACTOR3 and ARF4 Underlies the Wiry Leaf Syndrome  

Abstract

Interfering with small RNA production is a common strategy of plant viruses. A unique class of small RNAs that require microRNA and short interfering (siRNA) biogenesis for their production is termed trans-acting short interfering RNAs (tasiRNAs). Tomato (Solanum lycopersicum) wiry mutants represent a class of phenotype that mimics viral infection symptoms, including shoestring leaves that lack leaf blade expansion. Here, we show that four WIRY genes are involved in siRNA biogenesis, and in their corresponding mutants, levels of ta-siRNAs that regulate AUXIN RESPONSE FACTOR3 (ARF3) and ARF4 are reduced, while levels of their target ARFs are elevated. Reducing activity of both ARF3 and ARF4 can rescue the wiry leaf lamina, and increased activity of either can phenocopy wiry leaves. Thus, a failure to negatively regulate these ARFs underlies tomato shoestring leaves. Overexpression of these ARFs in Arabidopsis thaliana, tobacco (Nicotiana tabacum), and potato (Solanum tuberosum) failed to produce wiry leaves, suggesting that the dramatic response in tomato is exceptional. As negative regulation of orthologs of these ARFs by ta-siRNA is common to land plants, we propose that ta-siRNA levels serve as universal sensors for interference with small RNA biogenesis, and changes in their levels direct species-specific responses.

The ASYMMETRIC LEAVES Complex Employs Multiple Modes of Regulation to Affect Adaxial-Abaxial Patterning and Leaf Complexity

“…While links between adaxial-abaxial polarity and leaf complexity have been noted (Kim et al, 2003;Yifhar et al, 2012), our data indicate that altered adaxial-abaxial patterning is unlikely to drive this phenotype. Much like the examples described above, the increase in leaf complexity derives from the misregulation of their shared target ARF3 and the presence of additional factor(s) normally repressed by AS1-AS2.…”

“…Instead, we propose that ARF3, in conjunction with additional AS pathway targets, promotes organogenic potential at marginal auxin maxima, such that its prolonged activity results in elaboration of serrations into leaflets. Such a role for ARF3 may be evolutionarily conserved, as hypomorphic mutations in ta-siRNA biogenesis components in tomato (Solanum lycopersicum) can condition mild increases in leaf complexity (Yifhar et al, 2012). Thus, our analyses show that the shared repression of ARF3 by the AS and ta-siRNA pathways intersects with multiple additional AS1-AS2 targets to affect discrete nodes in leaf development.…”

Leaf development and morphogenesis

“…Mutations in several genes in the tomato tasiRNA pathway, which are negative regulators of ARF2, 3 and 4, were shown to underlie the tomato 'wiry' syndrome of very narrow leaves with reduced complexity (Lesley and Lesley, 1928;Yifhar et al, 2012). Interestingly, compromised tasiRNA pathway activity in M. truncatula led to a milder phenotype of increased leaf lobing with no effect on the number of leaflets (Zhou et al, 2013), whereas leaf development in Arabidopsis was unaffected (Hunter et al, 2006).…”

The Sussex signal: insights into leaf dorsiventrality

“…For instance, Antirrhinum phan mutants show clear adaxial-abaxial polarity phenotypes, as do tobacco and tomato plants harboring mutations in phan orthologs, but rough sheath2 from maize and as1 from Arabidopsis do not (Timmermans et al, 1999;Tsiantis et al, 1999;Byrne et al, 2000;Kim et al, 2003a;McHale and Koning, 2004). Likewise, mutations affecting tasiARF biogenesis condition a strong abaxializing phenotype in maize, rice and tomato, but cause only subtle polarity defects in Arabidopsis (Nogueira et al, 2007;Nagasaki et al, 2007;Chitwood et al, 2009;Douglas et al, 2010;Yifhar et al, 2012;Dotto et al, 2014). The latter difference may be explained, in part, by variation in the spatiotemporal expression of pathway components across species; tasiARF, for example, acts in the incipient primordium in maize and rice but during later stages in Arabidopsis leaf development (see Husbands et al, 2009).…”

“…5). For example, miR166, which accumulates on the abaxial side of the leaf, guides the cleavage of HD-ZIPIII transcripts, limiting the expression of these adaxial determinants to the top side of primordia (Juarez et al, 2004a; Conversely, tasiARF, which is generated through the specialized TAS3 trans-acting siRNA pathway (see Chapman and Carrington, 2007 for details on this small RNA pathway) limits expression of the ARF3 and ARF4 targets to a precisely defined domain on the bottom side of developing primordia (Nogueira et al, 2007;Nagasaki et al, 2007;Chitwood et al, 2009;Yifhar et al, 2012;Petsch et al, 2015). Interestingly, although the individual network components are highly conserved between angiosperms, the extent to which they contribute to adaxial or abaxial cell fate is not.…”