The evolution of quantitative traits in complex environments.

Species inhabit complex environments and respond to selection imposed by numerous abiotic and biotic conditions that vary in both space and time. Environmental heterogeneity strongly influences trait evolution and patterns of adaptive population differentiation. For example, heterogeneity can favor local adaptation, or can promote the evolution of plastic genotypes that alter their phenotypes based on the conditions they encounter. Different abiotic and biotic agents of selection can act synergistically to either accelerate or constrain trait evolution. The environmental context has profound effects on quantitative genetic parameters. For instance, heritabilities measured in controlled conditions often exceed those measured in the field; thus, laboratory experiments could overestimate the potential for a population to respond to selection. Nevertheless, most studies of the genetic basis of ecologically relevant traits are conducted in simplified laboratory environments, which do not reflect the complexity of nature. Here, we advocate for manipulative field experiments in the native ranges of plant species that differ in mating system, life-history strategy and growth form. Field studies are vital to evaluate the roles of disparate agents of selection, to elucidate the targets of selection and to develop a nuanced perspective on the evolution of quantitative traits. Quantitative genetics field studies will also shed light on the potential for natural populations to adapt to novel climates in highly fragmented landscapes. Drawing from our experience with the ecological model system Boechera (Brassicaceae), we discuss advancements possible through dedicated field studies, highlight future research directions and examine the challenges associated with field studies.

The calmodulin-binding transcription factor SIGNAL RESPONSIVE1 is a novel regulator of glucosinolate metabolism and herbivory tolerance in Arabidopsis.

The Arabidopsis Ca(2+)/calmodulin (CaM)-binding transcription factor SIGNAL RESPONSIVE1 (AtSR1/CAMTA3) was previously identified as a key negative regulator of plant immune responses. Here, we report a new role for AtSR1 as a critical component of plant defense against insect herbivory. Loss of AtSR1 function impairs tolerance to feeding by the generalist herbivore Trichoplusia ni as well as wound-induced jasmonate accumulation. The susceptibility of the atsr1 mutant is associated with decreased total glucosinolate (GS) levels. The two key herbivory deterrents, indol-3-ylmethyl (I3M) and 4-methylsulfinylbutyl (4MSOB), showed the most significant reductions in atsr1 plants. Further, changes in AtSR1 transcript levels led to altered expression of several genes involved in GS metabolism including IQD1, MYB51 and AtST5a. Overall, our results establish AtSR1 as an important component of plant resistance to insect herbivory as well as one of only three described proteins involved in Ca(2+)/CaM-dependent signaling to function in the regulation of GS metabolism, providing a novel avenue for future investigations of plant-insect interactions.

Analyses of in vivo interaction and mobility of two spliceosomal proteins using FRAP and BiFC.

U1-70K, a U1 snRNP-specific protein, and serine/arginine-rich (SR) proteins are components of the spliceosome and play critical roles in both constitutive and alternative pre-mRNA splicing. However, the mobility properties of U1-70K, its in vivo interaction with SR proteins, and the mobility of the U1-70K-SR protein complex have not been studied in any system. Here, we studied the in vivo interaction of U1-70K with an SR protein (SR45) and the mobility of the U1-70K/SR protein complex using bimolecular fluorescence complementation (BiFC) and fluorescence recovery after photobleaching (FRAP). Our results show that U1-70K exchanges between speckles and the nucleoplasmic pool very rapidly and that this exchange is sensitive to ongoing transcription and phosphorylation. BiFC analyses showed that U1-70K and SR45 interacted primarily in speckles and that this interaction is mediated by the RS1 or RS2 domain of SR45. FRAP analyses showed considerably slower recovery of the SR45/U1-70K complex than either protein alone indicating that SR45/U1-70K complexes remain in the speckles for a longer duration. Furthermore, FRAP analyses with SR45/U1-70K complex in the presence of inhibitors of phosphorylation did not reveal any significant change compared to control cells, suggesting that the mobility of the complex is not affected by the status of protein phosphorylation. These results indicate that U1-70K, like SR splicing factors, moves rapidly in the nucleus ensuring its availability at various sites of splicing. Furthermore, although it appears that U1-70K moves by diffusion its mobility is regulated by phosphorylation and transcription.

Ligand-dependent reduction in the membrane mobility of FLAGELLIN SENSITIVE2, an arabidopsis receptor-like kinase.

Arabidopsis Flagellin sensitive2 (FLS2) is a transmembrane leucine-rich repeat receptor-like kinase, which recognizes a conserved 22 amino acid peptide (flg22) of bacterial flagellin and activates downstream defense signaling pathways resulting in enhanced resistance against plant pathogens. The underlying mechanisms for the activation of FLS2 in the cell membrane, however, are not fully understood. Using fluorescence recovery after photobleaching (FRAP), we demonstrate that approximately 75% of the FLS2 in the plasma membrane diffuses laterally with a diffusion coefficient of 0.34 microm(2) s(-1), indicating that it moves rapidly. Further, we show that FLS2 is less mobile in the presence of flg22, suggesting its ligand-dependent confinement to microdomains or transient interaction with other less mobile membrane proteins. Using an in vivo bimolecular fluorescence complementation (BiFC) system and fluorescence resonance energy transfer (FRET), which reveals in vivo protein-protein interactions, we show that FLS2 does not homodimerize either constitutively or in the presence of flg22. Our data suggest that the reduced mobility of FLS2 after binding flg22 and its existence in monomeric form are important mechanistic features of FLS2 early signaling.

Slow desiccation leads to high-frequency shoot recovery from transformed somatic embryos of cotton (Gossypium hirsutum L. cv. Coker 310 FR).

In Agrobacterium-mediated genetic transformation of cotton (Gossypium hirsutum L. cv. Coker 310FR) the frequency at which somatic embryos were converted to plantlets was significantly improved by subjecting the embryos to slow physical desiccation. We used Agrobacterium strain GV3101 containing the binary vector pGSFR with the nos-nptII gene for in vitro selection and the 35S gus-int fragment as a reporter to optimize the transformation protocol. Although the concentration of kanamycin was reduced during embryogenesis and embryo maturation, even at the lower levels somatic embryos were predominantly abnormal, showing hypertrophy and reduced or fused cotyledons or poor radicle ends. A majority of these embryos (more than 75%) were beta-glucuronidase (GUS)-positive. Embryos with an abnormal appearance showed a very poor conversion to plantlets. However, these embryos, when subjected to slow physical desiccation followed by transfer to fresh medium, regenerated single or multiple shoots from the cotyledonary end. These shoots could be grafted on wild-type seedling stocks in vitro, which, following their transfer to soil, developed normally and set seeds. Regenerated plants tested positive for the transgene by Southern analysis. An overall scheme for the high-frequency production of cotton transgenics from both normal and abnormal appearing somatic embryos is presented.