Molecular basis for differential light responses in Arabidopsis stems and leaves

Plants optimize their growth and development in response to changes in light quality, quantity, and direction. In dark conditions, Arabidopsis seedlings elongate their hypocotyls while suppressing the expansion and growth of their cotyledons. Fast stem elongation in the dark can ensure that seedlings quickly emerge from underneath the surface of soil to reach light. Once they perceive light, Arabidopsis seedlings expand their cotyledons and gain the capacity to start photosynthesis while greatly reducing the elongation of their hypocotyls. The differential growth rates between cotyledons and hypocotyls in both dark and light are important for plants to grow properly in different light environments, yet the molecular mechanisms by which light, or a lack thereof, can differentially regulate the growth of stems and leaves are not understood. In PNAS, Sun et al. (1) uncover the key roles of a group of Small Auxin Up RNA (SAUR) genes in determining the differential growth between stems and leaves (Fig. 1).

Fig. 1.

Differential expression of lirSAURs is a key determinant for the different growth rates of cotyledons and hypocotyls under various light conditions. (A) High lirSAUR levels stimulate growth, and low lirSAUR expression represses growth. Dark-grown plants have elevated lirSAUR expression in the elongated hypocotyls (red) and decreased expression of lirSAURs in the closed cotyledons (light blue). In contrast, light-grown seedlings have expanded cotyledons and short hypocotyls, which correlate with high lirSAUR expression in cotyledons and low lirSAUR expression in hypocotyls. Plants grown under shade have long hypocotyls, extended petioles, and small cotyledons. It is not clear whether lirSAURs also play a role in the differential growth of plants grown under shade. (B) Some SAUR genes are differentially expressed in plants grown under shade. A 1-h shade treatment is sufficient to induce the expression of many SAUR genes. Microarray data were previously reported by Tao et al. (17), and are available from the Gene Expression Omnibus (GEO) database (accession no. GSE9816). Wc, constant white light.

The transition from dark-grown to light-grown, a process that mimics the emergence of a germinating seedling from underneath the soil, has been studied extensively (2, 3). Gene expression profiling experiments have been conducted to determine the differential gene expression patterns between dark-grown Arabidopsis seedlings and light-grown plants, between WT and photoreceptor mutants, and between plants grown under different light conditions (4–8). Thousands of genes are known to express differentially in response to different light conditions or to the transition from dark to light. Sun et al. (1) also used gene expression profiling to investigate how light can inhibit growth in hypocotyls and promote cell expansion in cotyledons at the same time. The study by Sun et al. (1) focused on shorter transition time periods compared with previous studies (1 or 6 h vs. 36 h) (1, 4, 6). More importantly, Sun et al. (1) conducted organ-specific gene expression profiling so that they could understand how light signals lead to opposite growth patterns in stems and leaves. They analyzed three groups of organ-specific light-responsive genes (OLRs) and found that hormone-related genes were highly enriched in the OLRs. Interestingly, about 40% (77 of 208) of the hormone-related OLRs are involved in auxin pathways, which have long been recognized for their roles in diverse processes of plant growth and development, especially in differential growth in gravitropic and phototropic responses (9). Sun et al. (1) chose the SAURs for further characterization because the SAURs were remarkably enriched in the auxin-related OLRs. Sun et al. (1) were particularly interested in the 32 SAUR genes that are induced in cotyledons and/or repressed in hypocotyls (lirSAURs) because there appeared to be a correlation between lirSAUR expression patterns and the observed differential growth in hypocotyls and cotyledons.

SAURs were initially identified by Thomas Guilfoyle’s group as early auxin-inducible genes, but their functions have remained mysterious until very recently (10, 11). It was recently reported that SAUR19 could physically interact with and inhibit the PP2C-D subfamily of type 2C protein phosphatases, which regulate the phosphorylation status of plasma membrane H⁺-ATPases in Arabidopsis (12, 13). Consequently, plants overexpressing SAUR19 have increased plasma membrane H⁺-ATPase activities and increased cell expansion and growth (12, 13). Therefore, Sun et al. (1) hypothesize that the lirSAURs, which are differentially expressed in cotyledons and hypocotyls (Fig. 1), may play key roles in regulating the differential growth in the two organs. They overexpressed SAUR14, SAUR50, and SAUR65, three representative lirSAURs, as GFP fusion proteins in Arabidopsis and found that all of the SAUR overexpression lines had longer hypocotyls. The SAUR50 and SAUR65 overexpression lines also have larger cotyledons than WT plants. Furthermore, the SAUR overexpression lines had opened cotyledons when grown in darkness for an extended period. In light, the SAUR overexpression lines also had longer hypocotyls and larger cotyledons than WT, suggesting that the SAUR genes can promote growth in both cotyledons and hypocotyls regardless of the light conditions. To test their hypothesis further, Sun et al. (1) generated saur50saur16 double mutants using clustered regulatory interspaced short palindromic repeats/Cas9 technology. The saur16saur50 seedlings were significantly smaller than WT plants grown under the same conditions. Like SAUR19, the lirSAURs also interact and inhibit the PP2C-D phosphatases (1), indicating that SAURs probably use a general mechanism to promote growth.

Because many SAURs are induced by auxin (10, 11), it is reasonable to hypothesize that light causes differential auxin distribution/biosynthesis. Consequently, differential SAUR expression occurs in different organs in response to changes in light environments. However, regulation of SAUR expression was more complex than just the involvement of auxin. Sun et al. (1) show that light treatment for 1 h did not result in obvious changes in auxin levels in cotyledons, but during the same time frame, lirSAURs are induced in cotyledons. Unexpectedly, auxin treatments for 1 h did not induce the expression of lirSAURs in cotyledons, but did induce the expression of SAURs in hypocotyls under the same conditions. Therefore, it appears that the light-induced decrease of auxin levels in hypocotyls is partially responsible for the inhibition of hypocotyl elongation by light, whereas other non–auxin-related processes might play a more predominant role in up-regulating lirSAURs in cotyledons.

Phytochrome interacting factors (PIFs) are transcription factors that play important roles in photomorphogenesis (14, 15). Interestingly, Sun et al. (1) discovered that PIF3 and PIF4 bind directly to the promoters of lirSAURs. More importantly, almost all of the lirSAURs are up-regulated in cotyledons and down-regulated in hypocotyls in pif mutants (pif1pif3pif4pif5, pifq) (15) in dark-grown seedlings. Furthermore, the short hypocotyl

Results from Sun et al. clearly establish lirSAURs as key players in light-mediated differential growth in stems and leaves.

phenotypes of dark-grown pifq were partially rescued by overexpression of lirSAURs, suggesting that lirSAURs are indeed downstream of the PIFs and that lirSAURs are partially responsible for light-regulated cell expansion in hypocotyls. It would be interesting to investigate whether knockout of some lirSAURs in pifq could rescue the cotyledon phenotypes of dark-grown pifq.

Results from Sun et al. (1) clearly establish lirSAURs as key players in light-mediated differential growth in stems and leaves. SAURs have the capacity to regulate cell expansion, and the observed differential expression patterns of lirSAURs in cotyledons and hypocotyls correlate well with the differential growth between cotyledons and hypocotyls (Fig. 1). It will be intriguing to investigate whether lirSAURs also play significant roles in other situations that result in differential growth between cotyledons and hypocotyls. For example, Arabidopsis seedlings grown under shade have elongated hypocotyls and reduced cotyledon/leaf expansion (16) (Fig. 1). It is well known that shade induces auxin biosynthesis and that auxin biosynthetic mutants are defective in respond to shade (17–19). Moreover, PIFs also play essential roles in the shade avoidance response, and PIF mutants are essentially insensitive to shade (19). Therefore, we reanalyzed previously published microarray data generated from plants grown under constant light and under shade (17) to determine whether the SAUR genes are differentially expressed. Interestingly, many of the SAUR genes are indeed induced by shade (Fig. 1), suggesting that the increased SAUR expression may also be responsible for hypocotyl elongation under shade. A caveat of this analysis is that the microarray data were generated using whole seedlings and we do not know whether the observed increase of SAUR expression was restricted to hypocotyls. Nevertheless, the correlations between lirSAURs, cell elongation, and differential growth between hypocotyls and cotyledons appear true for both dark- and light-grown seedlings. Whether the lirSAURs play a similar role in regulating differential growth between cotyledons and hypocotyls under shade can be experimentally tested by analyzing the SAUR overexpression lines and the saur16saur50 double mutants grown under shade conditions.