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HILI destabilizes microtubules by suppressing phosphorylation and Gigaxonin-mediated degradation of TBCB

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ABSTRACT

Human PIWIL2, aka HILI, is a member of PIWI protein family and overexpresses in various tumors. However, the underlying mechanisms of HILI in tumorigenesis remain largely unknown. TBCB has a critical role in regulating microtubule dynamics and is overexpressed in many cancers. Here we report that HILI inhibits Gigaxonin-mediated TBCB ubiquitination and degradation by interacting with TBCB, promoting the binding between HSP90 and TBCB, and suppressing the interaction between Gigaxonin and TBCB. Meanwhile, HILI can also reduce phosphorylation level of TBCB induced by PAK1. Our results showed that HILI suppresses microtubule polymerization and promotes cell proliferation, migration and invasion via TBCB for the first time, revealing a novel mechanism for HILI in tumorigenesis.

No MeSH data available.


HILI interacts with TBCB.(A) Endogenous interaction between HILI and TBCB. (B) Immunofluorescence assays showed that HILI and TBCB were mainly overlapped in cytoplasm. (C) Schematic of HILI deletion mutants. (D) Schematic of TBCB deletion mutants. (E) Interaction between TBCB and different MYC-tagged HILI mutants. (F) Interaction between HILI and different MYC-tagged TBCB mutants.
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f2: HILI interacts with TBCB.(A) Endogenous interaction between HILI and TBCB. (B) Immunofluorescence assays showed that HILI and TBCB were mainly overlapped in cytoplasm. (C) Schematic of HILI deletion mutants. (D) Schematic of TBCB deletion mutants. (E) Interaction between TBCB and different MYC-tagged HILI mutants. (F) Interaction between HILI and different MYC-tagged TBCB mutants.

Mentions: We have proved that HILI suppresses microtubule polymerization in a TBCB-dependent manner, so we next confirmed whether HILI can interact with TBCB in tumor cells. Subsequently coimmunoprecipitation assays were performed, showing that endogenous HILI and TBCB can interact with each other in HeLa cells (Fig. 2A), and then immunofluorescence assays showed that endogenous HILI and TBCB were mainly overlapped in cytoplasm in HeLa and HepG2 cells (Fig. 2B). To further identify the functional domains involved in the interaction between HILI and TBCB, we constructed their mutants respectively (Fig. 2C,D). Results showed that mutants of HILI lacking the PAZ domain such as D1 or D4 failed to interact with TBCB (Fig. 2E). These results suggested that PAZ domain plays an important role in the binding between HILI and TBCB. We next mapped the HILI binding site in TBCB. TBCB mutants lacking the UBL domain such as d2 failed to interact with HILI (Fig. 2F). These results suggested that the HILI binding site in TBCB is localized in the UBL functional domain.


HILI destabilizes microtubules by suppressing phosphorylation and Gigaxonin-mediated degradation of TBCB
HILI interacts with TBCB.(A) Endogenous interaction between HILI and TBCB. (B) Immunofluorescence assays showed that HILI and TBCB were mainly overlapped in cytoplasm. (C) Schematic of HILI deletion mutants. (D) Schematic of TBCB deletion mutants. (E) Interaction between TBCB and different MYC-tagged HILI mutants. (F) Interaction between HILI and different MYC-tagged TBCB mutants.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5385498&req=5

f2: HILI interacts with TBCB.(A) Endogenous interaction between HILI and TBCB. (B) Immunofluorescence assays showed that HILI and TBCB were mainly overlapped in cytoplasm. (C) Schematic of HILI deletion mutants. (D) Schematic of TBCB deletion mutants. (E) Interaction between TBCB and different MYC-tagged HILI mutants. (F) Interaction between HILI and different MYC-tagged TBCB mutants.
Mentions: We have proved that HILI suppresses microtubule polymerization in a TBCB-dependent manner, so we next confirmed whether HILI can interact with TBCB in tumor cells. Subsequently coimmunoprecipitation assays were performed, showing that endogenous HILI and TBCB can interact with each other in HeLa cells (Fig. 2A), and then immunofluorescence assays showed that endogenous HILI and TBCB were mainly overlapped in cytoplasm in HeLa and HepG2 cells (Fig. 2B). To further identify the functional domains involved in the interaction between HILI and TBCB, we constructed their mutants respectively (Fig. 2C,D). Results showed that mutants of HILI lacking the PAZ domain such as D1 or D4 failed to interact with TBCB (Fig. 2E). These results suggested that PAZ domain plays an important role in the binding between HILI and TBCB. We next mapped the HILI binding site in TBCB. TBCB mutants lacking the UBL domain such as d2 failed to interact with HILI (Fig. 2F). These results suggested that the HILI binding site in TBCB is localized in the UBL functional domain.

View Article: PubMed Central - PubMed

ABSTRACT

Human PIWIL2, aka HILI, is a member of PIWI protein family and overexpresses in various tumors. However, the underlying mechanisms of HILI in tumorigenesis remain largely unknown. TBCB has a critical role in regulating microtubule dynamics and is overexpressed in many cancers. Here we report that HILI inhibits Gigaxonin-mediated TBCB ubiquitination and degradation by interacting with TBCB, promoting the binding between HSP90 and TBCB, and suppressing the interaction between Gigaxonin and TBCB. Meanwhile, HILI can also reduce phosphorylation level of TBCB induced by PAK1. Our results showed that HILI suppresses microtubule polymerization and promotes cell proliferation, migration and invasion via TBCB for the first time, revealing a novel mechanism for HILI in tumorigenesis.

No MeSH data available.