Activation of TLRs mediated the NF-κB signaling pathway plays an important pathophysiological role in cardiac hypertrophy. Triad3A, a ubiquitin E3 ligase, has been reported to negatively regulate NF-κB activation pathway via promoting ubiquitination and degradation of TLR4 and TLR9 in innate immune cells. The role of Triad3A in cardiac hypertrophic development remains unknown. The present study investigated whether there is a link between Triad3A and TLR4 and TLR9 in pressure overload induced cardiac hypertrophy. We observed that Triad3A levels were markedly reduced following transverse aortic constriction (TAC) induced cardiac hypertrophy. Similarly, stimulation of neonatal rat cardiac myocytes (NRCMs) with angiotensin-II (Ang II) significantly decreased Triad3A expression. To determine the role of Triad3A in TAC-induced cardiac hypertrophy, we transduced the myocardium with adenovirus expressing Triad3A followed by induction of TAC. We observed that increased expression of Triad3A significantly attenuated cardiac hypertrophy and improved cardiac function. To investigate the mechanisms by which Triad3A attenuated cardiac hypertrophy, we examined the Triad3A E3 ubiquitination on TLR4 and TLR9. We found that Triad3A promoted TLR4 and TLR9 degradation through ubiquitination. Triad3A mediated TLR4 and TLR9 degradation resulted in suppression of NF-κB activation. Our data suggest that Triad3A plays a protective role in the development of cardiac hypertrophy, at least through catalyzing ubiquitination-mediated degradation of TLR4 and TLR9, thus negatively regulating NF-κB activation.

Tesis por compendio / [EN] DELLA proteins are plant-specific transcriptional regulators known to relay environmental information to the transcriptional networks to modulate growth and development accordingly. The view of DELLAs as signalling hubs is justified by two characteristics: first, they control the activity of a large number of transcriptional factors (TFs) and other transcriptional regulators through physical interaction; and, second, they are degraded by the 26S proteasome in response to the phytohormone gibberellin (GA), whose metabolism is very sensitive to environmental stimuli (e.g. light, temperature, salt stress). However, at least two observations indicate that this mechanistic framework is still incomplete: (i) warm temperature destabilizes the GA-insensitive DELLA rga-¿17, indicating that DELLAs cannot be destabilized only by changes in GA levels; and (ii) when found at the chromatin, DELLAs are localized not only in gene promoters, but also in gene bodies, suggesting that DELLAs may regulate transcription through interactions with proteins other than TFs. In this Thesis, we provide evidence that shows how a different E3 ubiquitin ligase controls the stability of DELLAs in a GA-independent manner, and how DELLAs regulate gene expression by directly interacting with the basal transcriptional machinery. In the first chapter, using a combination of genetic, physiological, and molecular approaches we demonstrate that DELLAs are targeted to proteolytic degradation by the E3 ubiquitin ligase CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1). We show that COP1 interacts with the DELLAs GAI and RGA in vitro and in vivo, and that it promotes their polyubiquitination. We propose that COP1 represents a major pathway to degrade DELLAs in response to shade or to warm temperature. In the second chapter, we describe the interaction between DELLAs and the transcription elongation complex Polymerase-Associated Factor 1 (Paf1c). We show that Paf1c is required for the genome-wide deposition of monoubiquitinated H2B (H2Bub), a mark necessary for the progression of RNA polymerase II (RNAPII), and that this function is largely dependent on the presence of DELLAs. Likewise, impaired Paf1c or DELLA function results in a similar alteration in the accumulation and distribution of RNAPII in the Paf1c-target genes. We propose that DELLAs would exert this action by modulating the recruitment of Paf1c to the chromatin. These two new mechanisms underscore the importance of DELLAs as a central node in the environmental signalling network and should be considered in any potential application of DELLAs as biotechnological targets. / [ES] Las proteínas DELLA son reguladores transcripcionales específicos de plantas que transmiten información ambiental a las redes transcripcionales que modulan el crecimiento y el desarrollo. La propuesta de que las DELLAs actúan como "hubs" en redes de señalización se justifica por dos razones: primero, controlan la actividad de un gran número de factores de transcripción (FTs) y otros reguladores transcripcionales mediante interacción física; y segundo, son degradadas por el proteosoma 26S en respuesta a la fitohormona giberelina (GA), cuyo metabolismo es muy sensible a los estímulos ambientales (p. ej. luz, temperatura, estrés salino). Sin embargo, al menos dos observaciones sugieren que la información relativa a estos mecanismos no es completa: (i) las temperaturas altas desestabilizan incluso a rga-¿17, una versión de DELLA insensible a GAs, lo que indica que la estabilidad de las DELLAs no depende sólo de cambios en los niveles de GAs; y (ii) cuando se encuentran en la cromatina, las DELLAs no solo se posicionan en los promotores, sino también a lo largo de las regiones codificantes, lo que sugiere que las DELLAs podrían regular la transcripción mediante interacciones con otras proteínas diferentes a FTs. En esta Tesis, proporcionamos evidencia sobre una nueva E3 ubicuitina ligasa que controla la estabilidad de las DELLAs de una manera independiente a las GAs, y cómo las DELLA regulan la expresión génica interaccionando directamente con la maquinaria de transcripción basal. En el primer capítulo, usando una combinación de aproximaciones genéticas, fisiológicas y moleculares, demostramos que las DELLAs son marcadas para su degradación proteolítica por la E3 ubicuitina ligasa CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1). Mostramos que COP1 interacciona al menos con las DELLAs GAI y RGA in vitro e in vivo, y que promueve su poliubicuitinación. Proponemos que COP1 representa una vía importante de degradación de DELLAs en respuesta a sombra y temperaturas altas. En el segundo capítulo, describimos la interacción entre las DELLAs y el complejo de elongación transcripcional Polymerase-Associated Factor 1 (Paf1c). Mostramos que, como en animales, Paf1c se requiere para la deposición a nivel genómico de la H2B monoubiquitinada (H2Bub), una marca necesaria para la progresión de la RNA polimerasa II (RNAPII), y que esta función depende en gran medida de la presencia de DELLAs. Asimismo, la reducción de la función de las DELLAs provoca defectos equivalentes a los de la pérdida de función de Paf1c en cuanto a la acumulación y distribución de la RNAPII en los genes diana de Paf1c. Proponemos que las DELLAs podrían por tanto regular la transcripción modulando el reclutamiento de Paf1c a la cromatina. Estos nuevos mecanismos inciden en la importancia de las DELLAs como nodos centrales en las redes de señalización al ambiente y podrían ser considerados como dianas biotecnológicas en aproximaciones futuras. / [CA] Les proteïnes DELLA són reguladors transcripcionals específics de les plantes conegudes per transmetre informació mediambiental a les xarxes transcripcionals per modular el creixement i desenvolupament. La visió actual de DELLAs com a "hubs" de senyalització es justifica per dues característiques: en primer lloc, controlen l'activitat d'un gran nombre de factors transcripcionals (FTs) i d'altres reguladors transcripcionals mitjançant la interacció física; i, en segon lloc, es degraden pel proteosoma 26S en resposta a la fitohormona giberel.lina (GA), el metabolisme de la qual és molt sensible als estímuls ambientals (per exemple, la llum, la temperatura, l'estrès salí). No obstant això, almenys dues observacions indiquen que aquest marc mecanicista encara és incomplet: (i) la temperatura càlida desestabilitza la DELLA rga-¿17, que és insensible a GAs, indicant que les DELLA no es desestabilitzen només per canvis en els nivells d'aquesta fitohormona; i (ii) quan es troben a la cromatina, les DELLA no només es localitzen en els promotors dels gens, sinó també a la regió que es transcriu, cosa que suggereix que poden regular la transcripció mitjançant interaccions amb proteïnes diferents de FTs. En aquesta tesi, proporcionem evidències que mostren com una ubiquitina E3 lligasa diferent controla l'estabilitat de les proteïnes DELLA de manera independent de GAs, i com les DELLA regulen l'expressió gènica interactuant directament amb la maquinària transcripcional basal. En el primer capítol, mitjançant una combinació d'enfocaments genètics, fisiològics i moleculars, demostrem que les DELLA s'envien a la degradació proteolítica mitjançant la ubiquitina E3 lligasa CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1). Mostrem que la COP1 interacciona amb els DELLA GAI i RGA in vitro i in vivo, i que afavoreix la seva poliubiquitinació. Proposem que COP1 representa la via principal per degradar les DELLA en resposta a l'ombra o a la temperatura càlida. Al segon capítol, es descriu la interacció entre les DELLA i el complex d'allargament de transcripció Polymerase Associated Factor 1 (Paf1c). Mostrem que Paf1c es requereix per a la deposició a tot el genoma de H2B monoubiquitinada (H2Bub), una marca necessària per a la progressió de l'ARN Polimerasa II (RNAPII), i que aquesta funció depèn en gran part de la presència de DELLA. De la mateixa manera, quan la funció Paf1c o DELLA està deteriorada, es produeix una alteració similar en l'acumulació i distribució de RNAPII als gens diana de Paf1c. Proposem que les DELLA realitzen aquesta acció modulant el reclutament de Paf1c a la cromatina. Aquests dos nous mecanismes posen de manifest la importància de les proteïnes DELLA com a node central de la xarxa de senyalització ambiental i haurien de ser considerats en qualsevol aplicació potencial de DELLA com a objectius biotecnològics. / La realización de esta Tesis Doctoral ha sido posible gracias a un contrato predoctoral para la formación de doctores (BES-2014-068868) y a la ayuda a la movilidad predoctoral para la realización de estancias breves (EEBB-I-16-11070) del Ministerio de Economía y Competitividad, y a una beca EMBO Short-Term (number 8047). / Blanco Touriñán, N. (2020). New mechanisms of DELLA protein regulation and activity in Arabidopsis [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/149477 / TESIS / Compendio

Omi/HtrA2 is a mitochondrial serine protease with a dual and opposite function depending on its subcellular localization. Most of the previous studies focused on Omi/HtrA2’s pro-apoptotic function when the protein is released to the cytoplasm. It is becoming apparent that the main function of Omi/HtrA2 is within the mitochondria, where it has a pro-survival role. However, its mechanism is still poorly understood. To this end, we used the yeast two-hybrid system to dissect the Omi/HtrA2 pathway by identifying novel interactors and substrates. Our studies revealed a novel function of Omi/HtrA2 in the regulation of a deubiquitinating (DUB) complex. In addition we found that Omi/HtrA2 participates in mitophagy by regulating Mulan E3 ubiquitin ligase, which recruits GABARAP (gamma-amino-butyric acid receptor-associated protein) to the mitochondria. Abro1 is the scaffold protein of the DUB complex known as BRISC (BRCC36 isopeptidase complex) that is specific for Lys-63 deubiquitination. This complex is similar to the BRCA-1 complex, a known and important player in DNA damage response. Using the yeast two-hybrid screen and a bait consisting of the unique carboxy-terminus of the Abro1 protein, we identified three transcription factors which are members of the activating protein 1 (AP-1) family, namely ATF4, ATF5 and JunD. The AP-1 family member ATF4 is ubiquitously expressed, like Abro1, and important in cell cycle regulation and survival, thus we further analyzed this interaction. Abro1’s interaction with ATF4 was specific and present only when cells are under cellular stress. When Abro1 protein level is increased it provides protection against stress-induced cell iv death, but interaction between Abro1 and ATF4 is necessary to achieve this protection. The significance of this interaction was the translocation of Abro1 from the cytoplasm to the nucleus. These results establish a new cytoprotective function of cytoplasmic Omi/HtrA2 as a regulator of the BRISC DUB complex. Under normal conditions Omi/HtrA2 is localized in the intermembrane space (IMS) of the mitochondria. We have recently identified that the mitochondrial Mulan E3 ubiquitin ligase is a substrate of Omi/HtrA2 protease. Mulan, along with MARCH5/MITOL and RNF185, are the only three mitochondrial E3 ubiquitin ligases identified thus far. The function of Mulan has been linked to cell growth, cell death and autophagy/mitophagy. In addition, we showed that Omi/HtrA2, through regulation of the Mulan protein level, controls mitophagy, especially during mitochondrial stress. To understand Mulan’s function and its control by Omi/HtrA2, we set out to identify E2 conjugating enzymes that form a complex with Mulan E3 ligase. We isolated four specific interacting E2’s, namely Ube2E2, Ube2E3, Ube2G2 and Ube2L3. To identify substrates for each unique Mulan-E2 complex we used fusion baits in a second yeast two-hybrid screening. One of the interactors isolated against the Mulan-Ube2E3 bait was the GABARAP protein, a member of the Atg8 (autophagy) family. The mammalian Atg8 family is composed of seven members that have been linked to important roles in autophagy/mitophagy. We characterized this interaction both in vitro and in vivo and its role in mitophagy. Our results suggest that Mulan participates in various pathways, depending on the nature of its partner E2 conjugating enzyme. In addition, we identified the pathway by which Mulan participates in mitophagy by recruiting GABARAP to the mitochondria. v I want to dedicate this hard work to the people who mean the most to me, the people who have been more than supportive in these past five years; they are my very small, but very loud and loving, family. My brother Raffaello has always been proud of me and always told his friends how his sister was a "doctor." My father, Alvaro, always told me I had to be better than him, a man who has a master’s degree in chemistry. Finally, but most importantly to me, I want to dedicate this work and degree to my mother, Maria Aparecida Troncon, a woman who is like no other. She has always supported me, and with small gestures like having snacks ready when I came to visit after work or coming with me to lab on the weekends so I would not be alone, she told me every day that she was proud of me. Every time she met someone she told them I was her doctor and she said it with the biggest smile on her face. Unfortunately she passed away on December 17th, 2012, just months shy of the completion of my degree. I remember how proud she was when I received my bachelor’s and I can only imagine the size of her smile when I walk across the stage this time as a Ph.D. I love you mom, you are my rock and my strength; without your constant support and dedication I would have not reached this point. I know you are smiling at me from above. Last but not least I want to dedicate this to the man who is my better-half. Thank you for all your love and support, Guillermo. I love you.

Post-translational modifications are chemical changes that occur to proteins after their synthesis, which are essential to their function and regulation. Ubiquitination is a post translation modification that serves key roles in the regulation of proteins. USP7 is a deubiquitinase that has several critical substrates important for human health and disease, including the cancer relevant proteins PTEN, p53, MDM2 and DMNT1. Most of these substrates have been identified by cell biology or proteomics experiments, but a detailed biochemical and structural analysis is lacking, likely due to the challenge of generating site-specific and stoichiometrically ubiquitinated proteins. Therefore, we leveraged our expertise in protein semi-synthesis to generate these ubiquitinated substrates to study USP7’s ability to recognize and hydrolyze the ubiquitin, which will reveal key details on how USP7 selects its substrates. In our investigation, we generated several mono-ubiquitinated peptides, that could be functionalized later to install on the protein, and assayed USP7’s ability to hydrolyze the ubiquitin. / 2026-03-06T00:00:00Z

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Le détournement de la machinerie cellulaire basé sur la dégradation par la voie du protéasome est une stratégie fréquemment retrouvée chez les virus afin d’optimiser leur réplication. Ainsi, le VIH-1 a développé toute une série de contremesures via ses protéines accessoires, vif et vpu notamment, afin de cibler les facteurs de restriction vers la voie du protéasome. La protéine accessoire Vpr est également associée à un complexe Cul4 E3 ubiquitin ligase mais est toujoursorphelin de sa cible. Nos travaux ont montré que la protéine CTIP2 est un acteur majeur impliqué dans la restriction de la réplication du VIH-1. Nous proposons de défendre la thèse selon laquelle la protéine CTIP2 est dégradée par la voie du protéasome en présence de la protéine vpr. Nous avons ainsi montré que l’expression de la protéine CTIP2 est plus forte en absence qu’en présence de la protéine vpr. Des expériences utilisant des inhibiteurs de la voie du protéasome sont en faveur d’une régulation de type post traductionnel. Par immunoprécipitation, nous avons montré que CTIP2 fait partie d’un complexe comprenant DDB1 et DCAF1 en présence et en absence de Vpr. Sa dégradation est prévenue en présence du mutant vpr (Q65R) qui n’interagit plus avec DCAF, et en présence d’un Knock Down de DCAF1par ailleurs, DCAF1 est associé avec CTIP2 inclus dans le complexe impliqué dans l’établissement de la latence du VIH-1 comprenant notamment HDAC1. Enfin, les protéines CTIP2, Vpr et DCAF colocalisent dans les noyaux des cellules microgliales. Nos résultats suggèrent fortement que la protéine Vpr favorise la dégradation du facteur CTIP2, qui est décrit comme un facteur restreignant l’infection par le VIH-1 dans les cellules microgliales, et ainsi favorise sa réplication. / Usurping the host ubiquitination proteasome system (UPS) to inactivate the undesirable host protein is a common viral strategy. HIV-1 proteins inactivate the detrimental host proteins by this system. In Microglial cells, CTIP2 represses both initial phase and late phase of HIV-1 gene transcription. As HIV-1 can still replicate in the presence of CTIP2, we postulated that it might inactivate CTIP2 by using Cul4 E3 ubiquitin ligase complex to resume its replication. We observed higher CTIP2 expressions in the absence of Vpr, with no effect on CTIP2 mRNA and proteasome inhibitor can block this degradation. Co-immunoprecipitation assays showed that CTIP2 is associated with DCAF1 and DDB1 in the absence and presence of Vpr. We showed that this degradation is prevented by the using Vpr mutant (Q65R) and by knock down of DCAF1. Finally, we observed the co-localization of CTIP2 with Cul4A-DCAF1-DDB1 complex even in the absence of Vpr, in microglial cells. Additionally, DCAF1 interacts with CTIP2-associated heterochromatin enzymes complex. Our results suggest that Vpr expression increases the turnover of CTIP2 in HIV-1 productively infected cells. By degrading CTIP2, HIV-1 counteracts CTIP2-mediated silencing of its expression and favors its replication.

Les cellules infectées de façon latente constituent de sérieux obstacles à l'éradication du VIH et à la guérison complète des patients. Nous avons précédemment rapporté que le facteur cellulaire CTIP2 joue un rôle clé dans l'établissement et dans la persistance de la latence du VIH dans les cellules de la microglie, principaux réservoirs du virus dans le cerveau. En recrutant des complexes enzymatiques au niveau du promoteur viral, CTIP2 inhibe l'expression des gènes en favorisant la compaction de la chromatine et défavorise la réactivation des réservoirs viraux grâce à son activité inhibitrice de l’activité kinase du complexe d’élongation pTEFb. Cependant, nous ne savons pas comment le VIH-1 contrecarre les effets répresseurs de CTIP2 dans des cellules permissives à son expression. Manipuler la machinerie cellulaire d'ubiquitination afin de cibler les protéines hôtes indésirables est une stratégie commune utilisée par les rétrovirus. Ici, nous postulons que la protéine auxiliaire Vpr pourrait favoriser la dégradation de CTIP2 via le complexe CUL4-DDB1-DCAF1 pour contrer ses effets sur la réplication du VIH-1. Nos précédents résultats ont montré que CTIP2 contribuait à la réponse antivirale cellulaire grâce à son activité répressive sur la transcription du VIH. Nous avons montré que l'expression de CTIP2 était induite par un traitement à l'interféron-α suggérant que ce facteur fait partie de la réponse cellulaire à des infections virales. Nous avons observé que la réplication du wt- mais pas du mutant délété pour vpr diminue l'expression de CTIP2 dans des cellules infectées de manières productives. L'expression de Vpr a été corrélée avec une dégradation de CTIP2 et une augmentation de la transcription des gènes du VIH-1. De plus, nous avons montré par des expériences d’immunoprécipitation et de FRET/FLIM que la protéine CTIP2 interagit avec DDB1, DCAF1 et Vpr afin d'induire la dégradation de CTIP2 par la voie du protéasome. Enfin, nous démontrons que DCAF1 est nécessaire à la dégradation de CTIP2 par Vpr dans les noyaux des cellules infectées. Nos résultats suggèrent ainsi que la protéine virale Vpr détourne la machinerie cellulaire et plus spécifiquement la voie de dégradation du protéasome afin d'induire la dégradation de CTIP2. En dégradant CTIP2, le VIH-1 contrecarre une réponse cellulaire anti-virale et favorise ainsi sa réplication. Notre travail a permis de mieux comprendre la nature des mécanismes mis en jeu par le VIH-1 afin de favoriser sa réplication dans les cellules microgliales. / Latently infected cells constitute major blocks to HIV-1 eradication and a functional cure of the patients. We have previously reported that the cellular co-factor CTIP2 plays a key role in the establishment and persistence of HIV latency in microglial cells, the main reservoirs of the virus in the brain. By recruiting large enzymatic complexes at the viral promoter, CTIP2 silences HIV-1 gene transcription and disfavors the viral reactivation from the reservoirs. However, nothing is known on how HIV-1 can counteract the effects of CTIP2 in permissively infected cells. Usurping the host ubiquitination machinery to target undesirable host proteins is a common strategy utilized by retroviruses. Here, we tend to postulate that HIV-1 Vpr may target CTIP2 by Cul4A-DDB1-DCAF1 complex to counteract its effects on HIV-1 replication. Our results showed that CTIP2 contributes to the cellular anti-viral response. We demonstrated that interferon treatments induce expression of CTIP2 suggesting that this factor may be part of the cellular response to viral infections. We observed that replication of wt- but not Vpr-deleted HIV-1 reduced CTIP2 expression in productively infected cells. Vpr expression was correlated with low levels of CTIP2 and increased levels HIV-1 gene transcription. In addition, we showed that CTIP2 interacts with DDB1, DCAF1 and HIV-1 Vpr in order to induce the degradation of CTIP2 via proteasome by coimmunoprecipitation and FRET experiments. Finally, the abrogation of Vpr binding to the DCAF1-CUL4-DDB1 complex prevented CTIP2 degradation. Our results suggest that Vpr engages the ubiquitination machinery to induce CTIP2 degradation. By degrading CTIP2, HIV-1 counteracts CTIP2-mediated silencing of its expression and thus favors viral replication. Our work has helped to understand the nature of the mechanisms involved in HIV-1 to foster its replication in microglial cells. These results allow us to consider new strategies toward a cure of the patients.

La signalisation des brassinostéroïdes (BR) est importante pour presque tous les aspects du développement des plantes, comme en témoigne le phénotype extrêmement nain et stérile des mutants défectueux du récepteur des brassinostéroïdes BRASINOSTEROID INSENSITIVE 1 (BRI1). De plus, il est un régulateur clé de la réponse des plantes à l'augmentation de la température ambiante (thermomorphogenèse) dans les parties aériennes de la plante, associé à la signalisation auxine et au facteur de transcription PHYTOCHROME INTERACTING FACTOR 4 (PIF4). Cependant, les rôles des mécanismes moléculaires de la thermomorphogenèse des racines restent insaisissables. Dans cette thèse, je décris en détail les mécanismes moléculaires conduisant à la thermomorphogenèse des racines des plantes exposées à une température ambiante élevée à la suite de la germination. Pour que les plantes allongent leur racine primaire à 26 ° C, par rapport à 21 ° C, elles régulent sélectivement la signalisation BR via la dégradation de BRI1 en fonction de la température. De manière surprenante, dans nos propres conditions, la signalisation auxine n’est pas nécessaire pour la thermomorphogenèse radiculaire, ce qui suggère une différence entre les réponses de thermomorphogenèse aérienne et racinaire. En utilisant une approche de mutagenèse dirigée, nous avons pu déterminer que la dégradation est déclenchée par une modification post-traductionnelle ciblant les lysines, probablement l’ubiquitination K63. Pour découvrir l’ubiquitine ligase E3 impliquée dans la dégradation de BRI1 induite par la température, nous avons effectué un criblage double hybride en levure en utilisant le domaine cytoplasmique de BRI1. Nous avons obtenu trois protéines candidates nommées DENSE AND ERECT PANICLE (DEP), qui se localisent de manière surprenante dans des microtubules corticaux (MTc) et sont apparues en même temps que la signalisation par le BR, suggérant un lien fonctionnel. L'interaction entre DEP1 et BRI1 a été confirmée par trois techniques différentes et, par conséquent, les mutants simples dep sont défectueux dans la perception de BR. D'un côté, ils sont hyposensibles à la réduction de la longueur de l'hypocotyle induite par le BR, mais de l'autre, ils sont hypersensibles au gravitropisme induit par le BR. Ces données suggèrent une interaction entre la signalisation par BR, la dynamique sous-cellulaire de BRI1 et les microtubules corticaux. Des recherches futures permettront de mieux comprendre l'importance biologique de l'interaction BRI1-MTc en général et de l'interaction BRI1-DEP1 en particulier. / Brassinosteroid (BR) signaling is important for nearly all aspects of plant development, as attested by the extremely dwarf and sterile phenotype of mutants defective in the brassinosteroid receptor BRASINOSTEROID INSENSITIVE 1 (BRI1). In addition, it is a key regulator of plant responses to increase in ambient temperature (thermomorphogenesis) in the above-ground parts of the plant together with auxin signaling and the transcription factor PHYTOCHROME INTERACTING FACTOR 4 (PIF4). However, the roles molecular mechanisms of root thermomorphogenesis remain elusive. In this thesis, I describe in great detail the molecular mechanisms leading to root thermomorphogenesis of plants exposed to elevated ambient temperature from germination. In order for plants to elongate their primary root at 26°C, compared to 21°C, they selectively downregulate BR signaling via the temperature-specific degradation of BRI1. Surprisingly, under our own conditions, auxin signaling is not required for root thermomorphogenesis, suggesting a difference between aerial and root thermomorphogenesis responses. Using a site-directed mutagenesis approach, we are able to pinpoint that the degradation is triggered by a post-translational modification targeting lysines, probably K63 ubiquitination. To find out the E3 ubiquitin ligase involved in the BRI1 temperature-induced degradation we carried out a yeast two hybrid screen using BRI1’s cytoplasmic domain. We obtained three candidate proteins named DENSE AND ERECT PANICLE (DEP) that surprisingly localize to cortical microtubules (cMTs) and arose at the same time as BR signaling, suggesting a functional link. The interaction between DEP1 and BRI1 was confirmed by three different techniques and, consequently dep single mutants are defective in BR percepton. On one hand, they are hyposensitive to the BR-induced reduction in hypocotyl length but on the other hand they are hypersensitive regarding BR-induced agravitropism. This data suggest an interplay between BR signaling, BRI1 subcellular dynamics and cortical microtubules. Future research will shed light on the biological significance of the BRI1-cMTs interaction in general and the BRI1-DEP1 interaction in particular.