Activator Gcn4 employs multiple segments of Med15/Gal11, including the KIX domain, to recruit mediator to target genes in vivo
Authors: Iness Jedidi
Ira Palmer
Hongfang Qiu
Sujoy Mukherjee
Christopher P Jaroniec
Paul T Wingfield
Joshua D Kaufman
Philippe S Nadaud
Stephen J Stahl
Alan G Hinnebusch
Fan Zhang
Journal: Journal of Biological Chemistry (2010)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: Mediator; RNA polymerase II; Med15/Gal11; Gcn4; recruitment
Affiliations: Laboratory of Gene Regulation and Development, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA.

Abstract:  Mediator is a multisubunit coactivator required for initiation by RNA polymerase II. The Mediator tail subdomain, containing Med15/Gal11, is a target of the activator Gcn4 in vivo, critical for recruitment of native Mediator or the Mediator tail subdomain present in sin4Delta cells. Although several Gal11 segments were previously shown to bind Gcn4 in vitro, the importance of these interactions for recruitment of Mediator and transcriptional activation by Gcn4 in cells was unknown. We show that interaction of Gcn4 with the Mediator tail in vitro and recruitment of this subcomplex and intact Mediator to the ARG1 promoter in vivo involve additive contributions from three different segments in the N terminus of Gal11. These include the KIX domain, which is a critical target of other activators, and a region that shares a conserved motif (B-box) with mammalian coactivator SRC-1, and we establish that B-box is a critical determinant of Mediator recruitment by Gcn4. We further demonstrate that Gcn4 binds to the Gal11 KIX domain directly and, by NMR chemical shift analysis combined with mutational studies, we identify the likely binding site for Gcn4 on the KIX surface. Gcn4 is distinctive in relying on comparable contributions from multiple segments of Gal11 for efficient recruitment of Mediator in vivo. 

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 *[right]* Fig 6b.  The tumor weights of mice with different treatments, including PBS (control), Dox (2 mg kg), Dox-GO (containing 2 mg kg of Dox), and ANG-Dox-GO (containing 2 mg kg of Dox), respectively, on 20 days . *[left]* Fig 5b from [Mu et al (2020)] . Surprising similarities. ![file] , 
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 *[left]* Fig 1A.  Figure 1. Characterization of angiopep-2 polypeptide-modified and doxorubicin-loaded graphene oxide (ANG-Dox-GO). (A) The appearance of commercial GO . *[right]* Fig 1A from [Wang et al (2020)] ,  Figure 1. Characterization of graphene oxide (GO). (A) Appearance of commercial GO dispersion . These appear to be the same bottle, with identical highlights, photographed in front of the same background, at slightly different angles. ![file] The descriptions are surprisingly similar, even accepting that there are only so many ways to describe the appearance of graphene suspension. Respectively,  The color of commercial GO varies from yellow to brown (Figure 1A). Under a AFM, ANG-Dox-GO was 4-5 nm in thickness and 100-400 nm in diameter (Figure 1B), suggesting the formation of single or several layers of GO nanosheets. In addition, the optical absorption spectra of ANG-Dox-GO showed absorption peaks at approximately 230 nm (Figure 1C).  The color of commercial GO dispersion varied from yellow to brown (Figure 1A). The optical absorption spectra of GO showed absorption peaks at approximately (Figure 1B). As determined by atomic force microscopy, GO was 3-4 nm in thickness and 100- 200 nm in diameter (Figure 1C), suggesting the formation of single or a few layers of GO nanosheets (Figure 1). , 
 #1 A subset of these tumors previously appeared as Fig 6B of [Zheng et al (2020)] . ![] UPDATE: Another sighting! Fig 6A of [Tian et al (2020)] . ![] ,