An N-terminal Nuclear Export Signal Regulates Trafficking and Aggregation of Huntingtin (Htt) Protein Exon 1*

Abstract

Background: Trafficking of huntingtin (Htt) fragments influences its toxicity. Results: A leucine-rich NES lies within the first 17 amino acids (N17) of Htt that controls subcellular localization and aggregation. Conclusion: The NES functions in cis and regulates the aggregation of Htt. Significance: This helps explain the mechanism of subcellular trafficking and aggregation of Htt fragments and may help elucidate molecular mechanisms of Htt toxicity.

Detection of huntingtin exon 1 phosphorylation by Phos-Tag SDS-PAGE: Predominant phosphorylation on threonine 3 and regulation by IKKβ

“…Although HEK293T cells are not a neuronal line, these cells are widely employed to model various huntingtin and mutant huntingtin characteristics including its phosphorylation status [4,12,13,15,18,23e25]. In order to study the effects of mutations on phosphorylation-prone residues on phenotypes known to be influenced by N17 phosphorylation such as subcellular localization and aggregation [5,10,12,13] and to enable correlations with phosphorylation status (results of these studies will be reported elsewhere), the huntingtin exon 1 constructs were originally designed to bear a C-terminal EGFP fusion (HTT-EX1-EGFP) enabling accurate quantification by imaging protocols. Therefore, the phosphorylation status of HTT-EX1-Q16-EGFP expressed in HEK293T cells was analyzed by Phos-Tag SDS-PAGE.…”

Lost in Transportation: Nucleocytoplasmic Transport Defects in ALS and Other Neurodegenerative Diseases

“…Some evidence suggests that reduced nuclear export of Htt may be one mechanism resulting in accumulation of the protein in the nucleus (Truant et al, 2007). The N-terminal 17 amino acids of Htt compose a consensus exportin-1–dependent nuclear export signal, which promotes shuttling between the cytoplasm and the nucleus (Maiuri et al, 2013; Zheng et al, 2013). Interestingly, the N-terminal Htt fragment directly interacts with nucleoprotein TPR (Figure 1), and this interaction is inhibited by the presence of a polyglutamine-expanded repeat and Htt aggregation, thus decreasing Htt export to the cytoplasm and increasing nuclear accumulation of mutant Htt (Cornett et al, 2005).…”

Integration-independent Transgenic Huntington Disease Fragment Mouse Models Reveal Distinct Phenotypes and Life Span in Vivo

“…The protein contains several known domains, including the polyQ domain, nuclear export signals (22)(23)(24), and proteolysis-susceptibility domains (Fig. 1A).…”