PTB/nPTB: master regulators of neuronal fate in mammals

Abstract

PTB was initially discovered as a polypyrimidine tract-binding protein (hence the name), which corresponds to a specific RNA-binding protein associated with heterogeneous ribonucleoprotein particle (hnRNP I). The PTB family consists of three members in mammalian genomes, with PTBP1 (PTB) expressed in most cell types, PTBP2 (also known as nPTB or brPTB) exclusively found in the nervous system, and PTBP3 (also known as ROD1) predominately detected in immune cells. During neural development, PTB is down-regulated, which induces nPTB, and the expression of both PTB and nPTB becomes diminished when neurons mature. This programed switch, which largely takes place at the splicing level, is critical for the development of the nervous system, with PTB playing a central role in neuronal induction and nPTB guarding neuronal maturation. Remarkably, sequential knockdown of PTB and nPTB has been found to be necessary and sufficient to convert non-neuronal cells to the neuronal lineage. These findings, coupled with exquisite understanding of the molecular circuits regulated by these RNA-binding proteins, establish a critical foundation for their future applications in regenerative medicine.

Keywords: Auto- and cross-regulation of alternative splicing; MicroRNA; Neuronal fate determination; Polypyrimidine tract-binding proteins.

Fig. 1

Domains and comparison of three PTB family members in the human genome. Each PTB paralog contains four RNA recognition motifs (RRMs) responsible for protein–RNA and protein–protein interactions, the latter of which is thought to mediate PTB dimerization or multimerization during splicing control

Fig. 2

Position-dependent effects of PTB on regulated splicing. Compiling PTB-binding events on PTB-dependent exon inclusion events (red) versus PTB-dependent exon skipping events (blue) suggests that PTB action on flanking constitutive splice sites enhances the inclusion of the alternative exon in the middle, while PTB binding on or around the alternative exon causes skipping of the exon

Fig. 3

Auto- and cross-regulation of PTB family members at the splicing level. A PTB binds the flanking intronic sequences of the alternative exon 11 to auto-regulate its own expression, as exclusion of this exon will trigger NMD. B PTB also represses the inclusion of the alternative exon 10, which is required to generate full-length mRNA for nPTB in non-neuronal cells. PTB down-regulation would thus induce this exon to produce functional nPTB during neuronal development. C PTB and nPTB appear to function in a redundant fashion to repress the inclusion of the alternative exon 2 in PTBP3/ROD1, but the functional significance of this regulated splicing event remains to be determined

Fig. 4

Modulation of microRNA targeting by PTB. A PTB binding directly competes with microRNA targeting on the microRNA responsive element (MRE). PTB depletion will thus promote microRNA targeting, thus enhancing microRNA-mediated post-transcriptional repression of gene expression. B PTB binding may also disrupt RNA secondary structure, which shields some MREs. PTB binding would expose such MREs for microRNA targeting, thereby enhancing post-transcriptional silencing of those genes

Fig. 5

Regulation of two consecutive regulatory loops for neuronal induction and maturation by PTB and nPTB. A PTB functions as a breaker to miR-124 targeting of REST. Once PTB down-regulation is initiated, miR-124 becomes more efficient in targeting REST, and reduced REST further de-represses miR-124. This loop is self-enforced by two mechanisms because PTB itself is a target for miR-124 and reduced PTB also results in the inclusion of exon 11, together causing progressive PTB down-regulation during neuronal induction. B Reduced PTB expression de-represses nPTB due to the inclusion of the alternative exon 10. During neuronal maturation, induced miR-9 targets nPTB, which somehow leads to the induction of mature neuronal-specific transcription factor Brn2, and activated Brn2 further induces miR-9. Thus, PTB and nPTB function as two separate gatekeepers for neuronal induction and maturation. The two regulated loops are efficiently connected in mouse cells, likely because of high-level induction of miR-124, which is also capable of targeting nPTB, but the two loops have to be separately activated in human cells to generate functional neurons