Crystal Structure of a Variant PAM2 Motif of LARP4B Bound to the MLLE Domain of PABPC1

Abstract

Eukaryotic cells determine the protein output of their genetic program by regulating mRNA transcription, localization, translation and turnover rates. This regulation is accomplished by an ensemble of RNA-binding proteins (RBPs) that bind to any given mRNA, thus forming mRNPs. Poly(A) binding proteins (PABPs) are prominent members of virtually all mRNPs that possess poly(A) tails. They serve as multifunctional scaffolds, allowing the recruitment of diverse factors containing a poly(A)-interacting motif (PAM) into mRNPs. We present the crystal structure of the variant PAM motif (termed PAM2w) in the N-terminal part of the positive translation factor LARP4B, which binds to the MLLE domain of the poly(A) binding protein C1 cytoplasmic 1 (PABPC1). The structural analysis, along with mutational studies in vitro and in vivo, uncovered a new mode of interaction between PAM2 motifs and MLLE domains.

A MademoiseLLE domain binding platform links the key RNA transporter to endosomes

“…However, mutating all four PAM2 motifs did not interfere with endosomal mRNA transport, although interaction with MLLE Pab1 was lost [43], confirming a potential redundancy. Consistently, mutations in PAM2 of human LARP4B did not interfere with the function of stress granule recruitment, suggesting additional factors in this case [45].…”

“…Structural analysis revealed a common mode of binding, where the Leu and particularly the Phe of the PAM2 consensus motif xxLNxxAxEFxP (S3A Fig) are interacting with helix 2 and 3 as well as helix 3 and 5 of MLLE domain, respectively [28,29]. Indeed, the interaction of MLLE with a hydrophobic amino acid is highly conserved, which in most cases is Phe with a known exception in the variant PAM2w motif of LARP4 and LARP4A, where Trp is found (S3A Fig) [28,44,45].…”

A MademoiseLLE domain binding platform links the key RNA transporter to endosomes

“…However, mutating all four PAM2 motifs did not interfere with endosomal mRNA transport, although interaction with MLLE Pab1 was lost (Jankowski et al ., 2019), confirming a potential redundancy. Consistently, mutations in PAM2 of human LARP4B did not interfere with the function of stress granule recruitment, suggesting additional factors in this case (Grimm et al ., 2020).…”

“…Consistently, mutations in PAM2 of human LARP4B did not interfere with the function of stress granule recruitment, suggesting additional factors in this case (Grimm et al, 2020).…”

“…Indeed, the interaction of MLLE with a hydrophobic amino acid is highly conserved, which in most cases is Phe with a known exception in the variant PAM2w motif of LARP4 and LARP4A, where Trp is found (Fig. EV3A; Grimm et al ., 2020; Yang et al ., 2011; Xie et al ., 2014).…”