The Dynein Light Chain Tctex-1 Has a Dynein-Independent Role in Actin Remodeling during Neurite Outgrowth

Abstract

Coordinated microtubule and microfilament changes are essential for the morphological development of neurons; however, little is know about the underlying molecular machinery linking these two cytoskeletal systems. Similarly, the indispensable role of RhoGTPase family proteins has been demonstrated, but it is unknown how their activities are specifically regulated in different neurites. In this paper, we show that the cytoplasmic dynein light chain Tctex-1 plays a key role in multiple steps of hippocampal neuron development, including initial neurite sprouting, axon specification, and later dendritic elaboration. The neuritogenic effects elicited by Tctex-1 are independent from its cargo adaptor role for dynein motor transport. Finally, our data suggest that the selective high level of Tctex-1 at the growth cone of growing axons drives fast neurite extension by modulating actin dynamics and also Rac1 activity.

Phosphorylation of DYNLT1 at Serine 82 Regulates Microtubule Stability and Mitochondrial Permeabilization in Hypoxia

“…DYNLT1, widely expressed in a number of tissues in various species and a component of dynein complex, fulfills cargo binding function (Belmont and Mitchison, 1996;Chuang et al, 2001), and plays a key role in multiple processes including endomembrane organization, trafficking, mitosis, and MT organization (Nagano et al, 1998;Palmer et al, 2011). DYNLT1 displays an independent cargo adaptor role for dynein motor transport apart from its other neuritogenic effects (Chuang et al, 2005). Although numerous reports have addressed dynein subunits, it is still unclear how they function with other molecules in the cytosol.…”

A dynein independent role of Tctex-1 at the kinetochore

“…Second, depletion of Tctex-1 does not affect dynein self-removal and the removal of the mitotic checkpoint proteins from attached kinetochores, implying that Tctex-1 is not functionally important for kinetochore-associated dynein. Third, a phosphomimetic Tctex-1T94E protein, which has been shown to be unable to incorporate into the dynein complex in transfected cells 19 and now by coimmunoprecipitation analysis using mitotic cell extracts, can rescue mitotic defects caused by the depletion of endogenous Tctex-1 proteins just like its nonphosphorylatable counterpart Tctex-1T94A that can associate with the dynein complex. And fourth, depletion of the Ndc80 complex, but not ZW10-dynein, at unattached kinetochores results in a significant loss of Tctex-1.…”

“…The Tctex-1 shRNA construct and rescue plasmids are as described previously. 19,20 Transfection was performed using HiPerfect (QIAGEN) based on the manufacturer's instructions (48 hr or 72 hr, as indicated in figures/figure legends), and cells transfected with shRNA were identified by the co-expressed HcRed. The mDia3 siRNA was purchased from QIAGEN.…”

“…19 Only the nonphosphorylatable Tctex-1-T94A protein, but not the phosphomimetic Tctex-1-T94E protein, is able to associate with the dynein complex. 19 In agreement, we confirmed that both Flagwild type Tctex-1 and Flag-Tctex-1-T94A, but not Flag-Tctex-1-T94E, were co-immunoprecipitated with endogenous dynein (DIC) in mitotic extracts (Fig. 4A).…”

“…For example, the LC8 family members of dynein light chains have been shown to interact, besides dynein, with a large number of proteins. 18 Similarly, the Tctex family member of Tctex-1 (or DYNLT1) has dynein-independent roles in actin remodeling during neurite outgrowth 19 and in ciliary resorption. 20, 21 We now show that Tctex-1 associates with unattached kinetochores and participates in stable microtubule attachment independent of cytoplasmic dynein.…”