Purification of an insect juvenile hormone receptor complex enables insights into its post-translational phosphorylation

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60077344%3A_____%2F21%3A00549785" target="_blank" >RIV/60077344:_____/21:00549785 - isvavai.cz</a>

Result on the web

<a href="https://www.sciencedirect.com/science/article/pii/S0021925821011935/pdfft?md5=ca6e964441ca194640b8b079e570c909&pid=1-s2.0-S0021925821011935-main.pdf" target="_blank" >https://www.sciencedirect.com/science/article/pii/S0021925821011935/pdfft?md5=ca6e964441ca194640b8b079e570c909&pid=1-s2.0-S0021925821011935-main.pdf</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.jbc.2021.101387" target="_blank" >10.1016/j.jbc.2021.101387</a>

Result language

angličtina

Original language name

Purification of an insect juvenile hormone receptor complex enables insights into its post-translational phosphorylation

Original language description

Juvenile hormone (JH) plays vital roles in insect reproduction, development, and in many aspects of physiology. JH primarily acts at the gene-regulatory level through interaction with an intracellular receptor (JH receptor [JHR]), a ligand-activated complex of transcription factors consisting of the JH-binding protein methoprene-tolerant (MET) and its partner taiman (TAI). Initial studies indicated significance of post-transcriptional phosphorylation, subunit assembly, and nucleocytoplasmic transport of JHR in JH signaling. However, our knowledge of JHR regulation at the protein level remains rudimentary, partly because of the difficulty of obtaining purified and functional JHR proteins. Here, we present a method for high-yield expression and purification of JHR complexes from two insect species, the beetle T. castaneum and the mosquito Aedes aegypti. Recombinant JHR subunits from each species were coexpressed in an insect cell line using a baculovirus system. MET–TAI complexes were purified through affinity chromatography and anion exchange columns to yield proteins capable of binding both the hormonal ligand (JH III) and DNA bearing cognate JH-response elements. We further examined the beetle JHR complex in greater detail. Biochemical analyses and MS confirmed that T. castaneum JHR was a 1:1 heterodimer consisting of MET and Taiman proteins, stabilized by the JHR agonist ligand methoprene. Phosphoproteomics uncovered multiple phosphorylation sites in the MET protein, some of which were induced by methoprene treatment. Finally, we report a functional bipartite nuclear localization signal, straddled by phosphorylated residues, within the disordered C-terminal region of MET. Our present characterization of the recombinant JHR is an initial step toward understanding JHR structure and function.

Czech name

—

Czech description

—

Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

—

OECD FORD branch

10605 - Developmental biology

Project

<a href="/en/project/GX20-05151X" target="_blank" >GX20-05151X: Uncovering of missing genetic components and new chemical regulators of juvenile hormone signaling</a><br>

Continuities

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Publication year

2021

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

Journal of Biological Chemistry

ISSN

0021-9258

e-ISSN

1083-351X

Volume of the periodical

297

Issue of the periodical within the volume

6

Country of publishing house

US - UNITED STATES

Number of pages

21

Pages from-to

101387

UT code for WoS article

000748084900001

EID of the result in the Scopus database

2-s2.0-85111949580