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Conjunction of epithelial cells into monolayer sheets implies the ability to migrate and to undergo apicobasal polarization. Both processes comprise reorganization of cytoskeletal elements and rearrangements of structural protein interactions. We modulated expression of tubulin tyrosin ligase (TTL), the enzyme that adds tyrosine to the carboxy terminus of detyrosinated α-tubulin, to study the role of tubulin detyrosination/-tyrosination in the orientation of cell motility and in epithelial morphogenesis. Oriented cell migration and the organization of focal adhesions significantly lose directionality with diminishing amounts of microtubules enriched in detyrosinated tubulin. On the other hand, increasing quantities of detyrosinated tubulin results in faster plus end elongation of microtubules in migrating and in polarized epithelial cells. These plus ends are decorated by the plus end binding protein 1 (EB1), which mediates interaction between microtubules enriched in detyrosinated tubulin and the integrin-ILK complex at focal adhesions. EB1 accumulates at the apical cell pole at the base of the primary cilium following apicobasal polarization. Polarized cells almost devoid of detyrosinated tubulin form stunted primary cilia and multiluminal cysts in 3D-matrices. We conclude that the balance between detyrosinated and tyrosinated tubulin alters microtubule dynamics, affects the orientation of focal adhesions and determines the organization of primary cilia on epithelial cells.

Antibody drug conjugates (ADCs), a promising class of cancer biopharmaceuticals , combine the specificity of therapeutic antibodies with the pharmacological potency of chemical, cytotoxic drugs. Ever since the first ADCs on the market, a plethora of novel ADC technologies has emerged, covering as diverse aspects as antibody engineering, chemical linker optimization and novel conjugation strategies, together aiming at constantly widening the therapeutic window for ADCs. This review primarily focuses on novel chemical and biotechnological strategies for the site-directed attachment of drugs that are currently validated for 2nd generation ADCs to promote conjugate homogeneity and overall stability. [ABSTRACT FROM AUTHOR]

Conjunction of epithelial cells into monolayer sheets implies the ability to migrate and to undergo apicobasal polarization. Both processes comprise reorganization of cytoskeletal elements and rearrangements of structural protein interactions. We modulated expression of tubulin tyrosin ligase (TTL), the enzyme that adds tyrosine to the carboxy terminus of detyrosinated α-tubulin, to study the role of tubulin detyrosination/-tyrosination in the orientation of cell motility and in epithelial morphogenesis. Oriented cell migration and the organization of focal adhesions significantly lose directionality with diminishing amounts of microtubules enriched in detyrosinated tubulin. On the other hand, increasing quantities of detyrosinated tubulin results in faster plus end elongation of microtubules in migrating and in polarized epithelial cells. These plus ends are decorated by the plus end binding protein 1 (EB1), which mediates interaction between microtubules enriched in detyrosinated tubulin and the integrin-ILK complex at focal adhesions. EB1 accumulates at the apical cell pole at the base of the primary cilium following apicobasal polarization. Polarized cells almost devoid of detyrosinated tubulin form stunted primary cilia and multiluminal cysts in 3D-matrices. We conclude that the balance between detyrosinated and tyrosinated tubulin alters microtubule dynamics, affects the orientation of focal adhesions and determines the organization of primary cilia on epithelial cells., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Müller, Gorek, Kamm and Jacob.)

Abstract: Tubulin partition between soluble and polymeric forms is tightly regulated in cells. Stathmin and tubulin tyrosine ligase (TTL) each form stable complexes with tubulin and inhibit tubulin polymerization. Here we explore the mutual relationship between these proteins in vitro and demonstrate that full-length stathmin and TTL compete for binding to tubulin and fail to make a stable tubulin:stathmin:TTL triple complex in solution. Moreover, stathmin depresses TTL tubulin tyrosination activity in vitro. These results suggest either that TTL and stathmin have a partially overlapping footprint on the tubulin dimer or that stathmin induces a tubulin conformation incompatible with stable TTL binding. [Copyright &y& Elsevier]

Tubulin, the building block of microtubules, is subject to chemically diverse and evolutionarily conserved post-translational modifications that mark microtubules for specific functions in the cell. Here we describe in vitro methods for generating homogenous acetylated, glutamylated, or tyrosinated tubulin and microtubules using recombinantly expressed and purified modification enzymes. The generation of differentially modified microtubules now enables a mechanistic dissection of the effects of tubulin post-translational modifications on the dynamics and mechanical properties of microtubules as well as the behavior of motors and microtubule-associated proteins., (© 2014 Elsevier Inc. All rights reserved.)