Lymphocyte p56^L32 is A RNA/DNA‐Binding Protein which Interacts with Conserved Elements of the Murine L32 Ribosomal Protein mRNA

Abstract

In previous studies of the ribosomal protein L32 mRNA, we demonstrated that a conserved polypyrimidine tract found in the 5'-untranslated region (5'-UTR) was required for translational regulation in vivo and that a 56-kDa protein (p56L32) from T-lymphocytes specifically interacts with this sequence [Kaspar, R. L., Kakegawa, T., Cranston, H., Morris, D. R. & White, M. W. (1992) J. Biol. Chem. 267, 508-514]. Here we show that p56L32 binding to the L32 5'-UTR is complex and requires other 5'-UTR RNA sequences in conjunction with the polypyrimidine tract. Deletion and site-directed mutagenesis studies revealed that binding of p56L32 to the L32 5'-UTR requires a second RNA element, GGUGGCUGCC, 15 nucleotides downstream from the polypyrimidine tract. In contrast, L32 RNA transcripts altered in this downstream element were good substrates for binding of the polypyrimidine binding proteins from HeLa nuclear extracts, indicating that these proteins have RNA-binding specificities distinct from p56L32. Competition analysis demonstrated that p56L32 will bind to DNA as well as RNA with identical sequence specificity and similar affinity. Single or double-stranded DNAs composed of the L32 5'-UTR sequences were found to specifically compete with L32 RNA transcripts for p56L32 binding. The L32 5'-UTR downstream element, GGUGGCUGCC, which is required for p56L32 binding, has previously been implicated as a transcriptional element of the L32 gene. The ability of p56L32 to bind this sequence as DNA or RNA suggests p56L32 may have a dual role in the regulation of ribosomal protein mRNA accumulation and translation.

TOP mRNAs are translationally inhibited by a titratable repressor in both wheat germ extract and reticulocyte lysate

“…Establishing the role of the 5′ TOP in the translational control of TOP mRNAs has led to the hypothesis that this motif might bind a specific translational trans ‐acting factor. Indeed, a cytoplasmic protein from mouse T‐lymphocytes was shown to specifically bind the first 34 nucleotides of mouse rpL32 mRNA [20,21]. Similarly, a sequence containing the first 52 nucleotides of Xenopus rpL1 mRNA directs binding of the La autoantigen to the oligopyrimidine tract and a cytoplasmic nucleic acids‐binding protein to sequences immediately downstream [22–24].…”

PTBP1 promotes hematopoietic stem cell maintenance and red blood cell development by ensuring sufficient availability of ribosomal constituents

“…Although binding of PTBP1 to these motifs has not previously been shown, our data suggest that PTBP1 associates with pyrimidine-rich sequences in 5 0 UTRs of selected transcripts, thereby affecting their protein-expression levels. Indeed, PTBP1 has previously been demonstrated to bind to a pyrimidine-rich sequence in the murine Rpl32 5 0 UTR (Severson et al, 1995), and a 57-kD protein (equals the size of PTBP1) was shown to bind the 5 0 UTR of Xenopus rpl1 mRNA (Cardinali et al, 1993). Moreover, PTBP1 has been reported to increase the translational efficiency of the FLT3mRNA in AML (Sun et al, 2019) and was also identified in a proteomics screen for proteins binding to 5 0 -cap structures of TOP transcripts following insulin-mediated stimulation of protein synthesis.…”

Μελέτη των όξινων ριβοσωμικών πρωτεϊνών και των γονιδίων τους στο έντομο Ceratitis capitata

“…Πράγματι, διαπιστώθηκε ότι μία κυτταροπλασματική πρωτείνη, περίπου 56 kDa, από μία σειρά Τ λεμφοκυττάρων έχει ικανότητα ειδικής πρόσδεσης στην 5 ' αμετάφραστη περιοχή του μηνύματος της rpL32 του ποντικού. Η πρόσδεση αυτή εξαρτάται από την ύπαρξη της 5 ' τελικής περιοχής των πυριμιδινών και από ένα δεύτερο δομικό στοιχείο, 15 νουκλεοτίδια δεξιά από το πρώτο (Kaspar et al, 1992;Severson et al, 1995). Παρόμοια, στον Xenopus βρέθηκε οτι, δύο πρωτείνες αλληλεπιδρούν ειδικά in vitro με την 5' αμετάφραστη περιοχή του μηνύματος της rpL4: η πρωτείνη Α, μεγέθους περίπου 57 kDa -ομόλογη της πρωτείνης που είχε χαρακτηριστεί στον ποντικό -η οποία προσδένεται στην 5' τελική περιοχή των πυριμιδινών, και η πρωτείνη C, μεγέθους περίπου 31 kDa, η οποία προσδένεται λίγα νουκλεοτίδια πιο δεξιά (Cardinali et al, 1993).…”