STAT5B-mediated Growth Hormone Signaling Is Organized by Highly Dynamic Microtubules in Hepatic Cells

Abstract

In the last decade, the notion that microtubules are critical to the spatial organization of signal transduction and contribute to the transmission of signals to downstream targets has been proposed. Because the STAT5B transduction and transcription factor is the major STAT protein activated by growth hormone stimulation in hepatocytes and is a crossroads between many signaling pathways, we studied the involvement of microtubules in STAT5B-mediated growth hormone signaling pathway in the highly differentiated and polarized WIF-B hepatic cell line. We showed that depolymerization of the microtubule network impaired STAT5B translocation to the nucleus upon growth hormone treatment. A significant amount of STAT5B binds to microtubules, while STAT5A and STAT3 are exclusively compartmentalized in the cytosol. Moreover, taxol-induced stabilization of microtubules released STAT5B from its binding, and we show that STAT5B binds specifically to the highly dynamic microtubules and is absent of the stable microtubule subpopulation. The specific involvement of dynamic microtubule subpopulation in growth hormone signaling pathway was confirmed by the inhibition of growth hormone-induced STAT5B nuclear translocation after stabilization of microtubules or specific disruption of highly dynamic microtubules. Upon growth hormone treatment, MT-bound STAT5B was rapidly released from microtubules by a dynein-dependent transport to the nucleus. Altogether, our findings indicate that the labile microtubule subpopulation specifically and dynamically organizes STAT5B-mediated growth hormone signaling in hepatic cells.Once bound to specific receptors, a large number of cytokines, growth factors, and hormones act through a sequence of steps allowing the rapid transport of information from the plasma membrane to subcellular targets. This information often results in the phosphorylation of transcription factors that subsequently migrate to the nucleus where they initiate specific gene transcription. How transcription factors are organized as molecular targets downstream of receptor activation and how they move rapidly through the cytoplasm from their activation site to the nucleus often remains unclear. The notion that microtubules (MTs) 1 might be involved in the cytoplasmic activation and/or trafficking of various transcription factors and protein kinases has emerged, as many signal transduction molecules, including transcription factors and protein kinases, have been shown to interact with MTs (1). For example, the glucocorticoid receptor was shown to colocalize with the MT network in mammalian cells (2, 3). The tumor suppressor protein p53 also uses MT tracks to migrate to the nucleus (4). Various protein kinases such as the MAP kinase kinase kinase MLK2 (5) or the MAP kinases ERK1 and ERK2 (6) were also shown to interact or to colocalize with MTs or with microtubule-associated proteins (7).In this study, we investigated the putative involvement of the MT network in GH signaling in the highly differentiated WIF-B hepatic cell line (8, 9...

Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL-7 Receptor Signalosome in Human CD4 T-cells

“…Induction of cytoskeleton structuring has been described with many cytokines and chemokines but mostly over longer time spans. Pioneering studies on prolactin receptor (35), growth hormone receptor (36), and epidermal growth factor receptor (37) have suggested that machineries carried by the cytoskeleton might be involved in STAT5 and pSTAT5 distribution and proposed that rapid organization was involved. Here, in our study, we used super-resolution optical microscopy to describe in detail how radial microtubules are formed within a few min of IL-7 activation, bridging plasma and nuclear membranes across the narrow cytoplasm of CD4 T-lymphocytes.…”

Requirement for the Dynein Light Chain km23-1 in a Smad2-dependent Transforming Growth Factor-β Signaling Pathway

“…For example, disruption of dynein motor activity by overexpressing dynamitin impaired the accumulation of p53 in the nucleus following DNA damage (71). In addition, overexpression of p50/dynamitin impaired the nuclear accumulation of STAT5B after growth hormone induction (72), as well as the nuclear translocation of the glucocorticoid receptors after ligand stimulation (73). Because our results have shown that overexpression of dynamitin blocked Smad2 nuclear translocation after TGF␤ treatment, dynein-dependent intracellular events also appear to be required for TGF␤/Smad2 downstream effects.…”

Starvation-induced Hyperacetylation of Tubulin Is Required for the Stimulation of Autophagy by Nutrient Deprivation

“…MT protein fractionation was then validated by checking that the signaling and transcription factor STAT5B bound specifically to the labile MT fraction ( Fig. 2A) and that the latter was not contaminated with soluble proteins, as STAT5A remained exclusively in the cytosol (36). The stable MT fraction was enriched in modified tubulin as shown by the distribution of the acetylated and detyrosinated tubulins.…”

“…To test this hypothesis, we performed cell fractionation experiments in which we purified the proteins associated with labile MTs and with stable MTs (36). Cells were permeabilized using low saponin concentrations to avoid artifactual LC3 aggregation (supplemental Fig.…”