The DNA-Binding High-Mobility Group Box Domain of Sox Family Proteins Directly Interacts with RNA In Vitro

Abstract

There is a growing body of evidence that a substantial number of protein domains identified as DNA-binding also interact with RNA to regulate biological processes. Several recent studies have revealed that the Sox2 transcription factor binds RNA through its high-mobility group box (HMGB) domain in vitro and in vivo. A high degree of conservation of this domain among members of the Sox family of transcription factors suggests that RNA-binding activity may be a general feature of these proteins. To address this hypothesis, we examined a subset of HMGB domains from human Sox family of proteins for their ability to bind both DNA and RNA in vitro. We observed selective, high-affinity interactions between Sox family HMGB domains and various model RNA elements, including a four-way junction RNA, a hairpin RNA with an internal bulge, G-quadruplex RNA, and a fragment of long noncoding RNA ES2, which is known to directly interact with Sox2. Importantly, the HMGB domains bind these RNA ligands significantly tighter than nonconsensus dsDNA and in some cases with affinities rivaling those of their consensus dsDNA sequences. These data suggest that RNA binding is a conserved feature of the Sox family of transcription factors with the potential to modulate unappreciated biological functions.

The DNA binding high mobility group box protein family functionally binds RNA

“…Additionally, the more well-studied HMGBs are more likely to have been investigated as an RBP whereas RNA binding potential may not yet have been explored for the lesser-studied HMGBs. The likelihood for other HMGB-RNA interactions is reinforced by prediction software which suggests the majority of Li, Zhu, et al, 2021) Sox1 (Hou et al, 2020) Unknown Sox2 (Hamilton et al, 2022;Holmes et al, 2020), (Cajigas et al, 2021;Hou et al, 2020;Tung et al, 2010) (Bai et al, 2022;Du et al, 2021;Guo et al, 2018;Jiang et al, 2018;Jing et al, 2020;Q. Li, Sun, et al, 2021;Ng et al, 2013;Ng et al, 2012;Samudyata, Amaral, et al, 2019;Stamou et al, 2020;Zhou et al, 2022;Zhuang et al, 2015) Sox4 Unknown (Chen et al, 2020) Sox6 (Hamilton et al, 2022) Unknown Sox9 (Hamilton et al, 2022) (Girardot et al, 2018;Pei et al, 2021;Penrad-Mobayed et al, 2018) Sox10 Unknown (Stamou et al, 2020) Sox11 (Hamilton et al, 2022) Unknown Sox13 (Yan et al, 2021) Unknown Sox15 (Hamilton et al, 2022) Unknown Sox17 (Hamilton et al, 2022) Unknown Sox21 (Hamilton et al, 2022) Unknown Sox30 (Hamilton et al, 2022) Unknown SRY (Hamilton et al, 2022) Unknown TCF7 (Lee & Jeong, 2004;Park et al, 2005) Unknown TFAM (Baltz et al, 2012;Brown et al, ...…”

“…In vitro, Sox2 interacts with a variety of RNA ligands with this interaction likely mediated by the HMGB‐DBD (Table 2; Cajigas et al, 2021; Hamilton et al, 2022; Holmes et al, 2020; Hou et al, 2020; Tung et al, 2010). Sox2 HMGB‐DBD interacts with G‐quadruplex RNA (1 nM), four‐way junction RNA (5 nM), fragment of lncRNA ES2 (3 nM), and varied stem‐loop RNAs (~10–100 nM; Hamilton et al, 2022; Holmes et al, 2020).…”

“…In vitro, Sox2 interacts with a variety of RNA ligands with this interaction likely mediated by the HMGB‐DBD (Table 2; Cajigas et al, 2021; Hamilton et al, 2022; Holmes et al, 2020; Hou et al, 2020; Tung et al, 2010). Sox2 HMGB‐DBD interacts with G‐quadruplex RNA (1 nM), four‐way junction RNA (5 nM), fragment of lncRNA ES2 (3 nM), and varied stem‐loop RNAs (~10–100 nM; Hamilton et al, 2022; Holmes et al, 2020). Sox2–HMGB‐DBD's affinity for RNA is comparable to dsDNA containing the ChIP‐validated Sox2 consensus motif ( K D = ~1 nM), but much tighter than to dsDNA without the Sox consensus motif ( K D = ~600 nM; Hamilton et al, 2022; Holmes et al, 2020).…”

“…Sox2 HMGB‐DBD interacts with G‐quadruplex RNA (1 nM), four‐way junction RNA (5 nM), fragment of lncRNA ES2 (3 nM), and varied stem‐loop RNAs (~10–100 nM; Hamilton et al, 2022; Holmes et al, 2020). Sox2–HMGB‐DBD's affinity for RNA is comparable to dsDNA containing the ChIP‐validated Sox2 consensus motif ( K D = ~1 nM), but much tighter than to dsDNA without the Sox consensus motif ( K D = ~600 nM; Hamilton et al, 2022; Holmes et al, 2020).…”

SOX1 acts as a tumor hypnotist rendering nasopharyngeal carcinoma cells refractory to chemotherapy

“…7C). SOX family proteins share a conserved high-mobility group (HMG) domain, binding to the same core consensus DNA motif (5'-TTGT) [23]. We analyzed de novo and known motifs in SOX1 peaks from the GEO database (Fig.…”