The microtubule cytoskeleton network in thyroid development and physiology and platelet functions – MITHYPLA
The microtubule cytoskeleton network in thyroid and platelet functions
β1-tubulin, a key component of the microtubule cytoskeleton, plays an essential role in platelet production and morphology. Recently, its involvement in thyroid development and function has also been demonstrated. We propose to investigate its role in platelet functions, which remains poorly understood. Furthermore, we hypothesize that post-translational modifications of microtubules also play a role in both the thyroid and platelets.
The role of the microtubule cytoskeleton network and its components in thyroid development and physiology and platelet functions
We have identified mutations in β1-tubulin (TUBB1) in patients with congenital hypothyroidism (CH) and macroplatelets. Our aim is to gain a better understanding of the role of TUBB1 and other β- and α-tubulin isoforms in thyroid development and physiology, as well as platelet function. Objectives - Task 1 : Decipher the role of TUBB1 mutations in thyroid development and platelet activation To ascertain the impact of TUBB1 mutations on thyroid development (P1) and platelet activation (P2). The specific functions of TUBB1 with known and newly discovered mutations in patients with CH will be deciphered. - Task 2 : Identify the role of MT network components (tubulin, MT interactors, tubulin modifiers) in thyroid and in platelets: To examine transgenic mice that have alterations in the post-translational modifications (PTMs) of tubulins. - Task 3 : Determine the role of MT in thyroid hormone secretion The aim is to gain a better understanding of CH and the role of an emerging regulatory mechanism of microtubules, the 'tubulin code', in the organization of the cytoskeleton and its regulation within this rare disease. This work will help to define the management of CH patients who are likely to be at increased risk of thrombosis. This research will provide a valuable framework for understanding the functions of MT in thyroid physiology and platelet function, contributing to a more comprehensive understanding of the mechanisms underlying various human diseases, including endocrine disorders.
Task1:
Cellular models: The eight TUBB1 mutations identified in patients with HC and macroplatelets were inserted into a lentiviral vector containing the human TUBB1 cDNA. Following lentiviral transduction, the effect of TUBB1 protein mutations will be evaluated on thyrocyte proliferation and migration using the Incucyte system.
In addition, the HEL cell line, which exhibits megakaryocytic characteristics, will be transduced to enable expression of the TUBB1-P160L variant, thereby allowing the study of the mutation’s impact on microtubule organization.
Murine model: In a mouse model carrying the p.P160L-Tubb1 mutation, thyroid and hemostatic phenotypes are being investigated.
Task 2:
Quantitative PCR (qPCR) for tubulins and enzymes involved in tubulin PTMs are performed on human and murine thyroid tissue at various stages of development and in adulthood.
qPCR and immunoblot analyses of the different tubulin isoforms expressed during platelet biogenesis were performed.
Task 3 :
To decipher the role of tubulins and PTM enzymes in thyroid hormone (TH) secretion, murine embryonic thyroid cultures were established. Tubulins and the genes encoding PTM enzymes were analyzed throughout this culture period in order to identify the most relevant expression profiles during thyroid differentiation.
Cellular and transgenic models have enabled to unravel the mechanism of platelet hyperactivation caused by the TUBB1-P160L mutation.
Furthermore, research has identified two new genes that encode PTM enzymes and are involved in thyroid development and function. The involvement of these genes highlights the significance of a finely regulated microtubule network for normal thyroid development and normal TH (thyroid hormone) secretion. Any disruption would lead to CH. Consequently, mutations in these genes represent a new therapeutic target for patients with CH.
Our project has opened up new avenues for understanding congenital hypothyroidism and has highlighted the involvement of an emerging microtubule regulatory mechanism, the ‘tubulin code’, in the organization of the cytoskeleton and its regulation in this rare disease. New genes encoding microtubule components have been identified as being involved in the development and secretion of thyroid hormones. Furthermore, the platelet activation mechanism involving B1-tubulin has been identified.
These new genes and mechanisms will be investigated and open up new research projects to better understand their role in the thyroid and platelets.
Few genes are involved in thyroid dysgenesis (TD) causing congenital hypothyroidism (CH) that occurs in about 1:3000 newborns. Recently, we identified a novel gene, TUBB1, involved in CH due to TD. TUBB1 encodes for a ß-tubulin isotype – one of the basic building blocks of microtubules (MTs) and strongly represented in platelets. We have shown that defects in MTs affected thyroid development and platelets activation. We hypothesize that other components of cytoskeleton, such as other tubulin isoforms, their post-translational modifications and their regulatory proteins could also disturb thyroid development, physiology and platelet function. Our project will decipher this hypothesis by analyzing the MT organization and their mechanisms in thyroid development, thyroid hormone secretion and platelet function in already established in vivo and in vitro models.
Project coordination
Michel POLAK (Institut Cochin)
The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.
Partnership
HITh Hémostase, Inflammation, Thrombose
BPPS BIOLOGIE ET PHARMACOLOGIE DES PLAQUETTES SANGUINES : HÉMOSTASES, THROMBOSE, TRANSFUSION (U 1255)
INSERM Institut Cochin
IC Institut Curie
Help of the ANR 583,574 euros
Beginning and duration of the scientific project:
January 2022
- 36 Months
