Investigation
Identification of new key proteins in the response to stress using translational and proteomics techniques
It has been widely reported that translation regulation plays an important role during plant response to environmental cues, allowing the translation of specific mRNAs that are crucial for the response. Based on this, our lab has implemented different techniques (such as super-resolution ribosome profiling, Ribo-seq analysis, polysomal profiling, etc.) to allow the identification of those mRNAs that are translationally regulated (and so, highly susceptible to play an important function) during specific stages of development and in response to environmental stresses.
As an example of these approaches, in the recent past, we carried out a high-throughput translatome analysis in Arabidopsis seedlings subjected to a moderate heat stress (38°C 1 h) (Yangüez et al., 2013). From this study, we identified different mRNAs that were selectively translated under heat stress conditions (Figure 1).
In addition, and with the same objective, we did a comparative proteomics analysis of plants grown under control conditions, during a heat acclimation event and at the early stages of the recovery periods from severe heat treatments that selectively lead plants to either survival or death. This quantitative proteomics analysis allowed us to unravel new proteins specifically involved in thermotolerance (Echevarría-Zomeño et al., 2016) (Figure 2).
The combined analysis of these translatomic and proteomic approaches retrieved different new potential regulators of plant adaptation to heat stress, which are being analyzed in the lab as part of our research activity.
Using the similar techniques we are actively identifying new novel regulators of plant response to different biotic and abiotic stresses.
Study of the role of different chaperones and co-chaperones in protein folding in response to different stresses
The previously described approaches (Yángüez et al., 2013 and Echevarría et al., 2016) allowed the identification of AtHOP3 as a potential regulator of thermotolerance in plants.
HOPs (HSP70-HSP90 organizing proteins) are a highly conserved family of co-chaperones, whose role in adaptation to stress was not fully evaluated in plants. In a recent study (Fernández-Bautista et al., 2017), we showed that HOP3 is induced during the unfolded protein response (UPR), interacts with the endoplasmic reticulum (ER)-resident protein BiP and plays an essential role during ER stress in plants. (Figure 3).
In addition, in a different report (Fernández-Bautista et al 2018), we described, for the first time in plants, that the three members of the AtHOP family act redundantly to promote long term acquired thermotolerance (LAT) in Arabidopsis. Our results demonstrated that HOP proteins are involved in two important processes associated to LAT, the full establishment of the transcriptional response during the acclimation period and the quality control maintenance during severe heat conditions. These data revealed that HOP family modulates the plant capacity to acclimate to high temperatures for long periods (Figure 4).
Recent results obtained in the lab suggest that the role of HOPs is not only circumvented to these two processes, but that HOPs are master hubs in the plant response to different stresses and during specific developmental programs. In this sense, one of our current interests is the identification of the possible targets of HOP and the analysis of how this interaction modifies their activity during the studied processes.
Identification of specific mechanisms of regulation of translation involved in plant adaptation to the environment
Translation regulation takes place mainly at the initiation step. In metazoans, one of the main mechanisms for translation regulation is the control of eIF4E activity by interaction with other proteins (known as 4E-BPs and 4E-binding partners). Despite the broad function of these eIF4E interacting proteins in the regulation of general and specific inhibition of translation in other eukaryotes, no orthologues have been found in plants (Sesma et al., 2017). In our lab, we have identified a novel translation regulator called CERES. CERES interacts with the eIF4E and regulates its function. However, in contrast to most of the metazoan eIF4-interacting proteins, which inhibit protein translation under stress and energy-limiting conditions, CERES favors the synthesis of general and specific proteins at precise time frames during the light cycle in plants, when their energy conditions (determined by photosynthesis) are optimal (Toribio et al., 2019) (Figure 5).
Following this work, we are currently focused on analyzing the role of CERES in other processes and in the deep characterization of its mechanism of action. In addition, we are also working in the characterization of other possible translational regulators in response to stress.
