Exploring the putative role of PRDM1 and PRDM2 transcripts as mediators of T lymphocyte activation

Abstract

Background T cell activation and programming from their naïve/resting state, characterized by widespread modifications in chromatin accessibility triggering extensive changes in transcriptional programs, is orchestrated by several cytokines and transcription regulators. PRDM1 and PRDM2 encode for proteins with PR/SET and zinc finger domains that control several biological processes, including cell differentiation, through epigenetic regulation of gene expression. Different transcripts leading to main protein isoforms with (PR +) or without (PR-) the PR/SET domain have been described. Although many studies have established the critical PRDM1 role in hematopoietic cell differentiation, maintenance and/or function, the single transcript contribution has not been investigated before. Otherwise, very few evidence is currently available on PRDM2. Here, we aimed to analyze the role of PRDM1 and PRDM2 different transcripts as mediators of T lymphocyte activation. Methods We analyzed the transcription signature of the main variants from PRDM1 (BLIMP1a and BLIMP1b) and PRDM2 (RIZ1 and RIZ2) genes, in human T lymphocytes and Jurkat cells overexpressing PRDM2 cDNAs following activation through different signals. Results T lymphocyte activation induced an early increase of RIZ2 and RIZ1 followed by BLIMP1b increase and finally by BLIMP1a increase. The “first” and the “second” signals shifted the balance towards the PR- forms for both genes. Interestingly, the PI3K signaling pathway modulated the RIZ1/RIZ2 ratio in favor of RIZ1 while the balance versus RIZ2 was promoted by MAPK pathway. Cytokines mediating different Jak/Stat signaling pathways (third signal) early modulated the expression of PRDM1 and PRDM2 and the relationship of their different transcripts confirming the early increase of the PR- transcripts. Different responses of T cell subpopulations were also observed. Jurkat cells showed that the acute transient RIZ2 increase promoted the balancing of PRDM1 forms towards BLIMP1b. The stable forced expression of RIZ1 or RIZ2 induced a significant variation in the expression of key transcription factors involved in T lymphocyte differentiation. The BLIMP1a/b balance shifted in favor of BLIMP1a in RIZ1-overexpressing cells and of BLIMP1b in RIZ2-overexpressing cells. Conclusions This study provides the first characterization of PRDM2 in T-lymphocyte activation/differentiation and novel insights on PRDM1 and PRDM2 transcription regulation during initial activation phases.

RIZ2 at the crossroad of the EGF/EGFR signaling in colorectal cancer

“…We have not used this approach here since it is still difficult to selectively silence RIZ2 without targeting, at least in part, also RIZ1 transcript [ 6 , 34 ]. In this context, one possibility could be to exploit the preferential presence of slightly different 3’ tails in RIZ1 and RIZ2 transcripts, as we recently described in lymphocytes [ 4 ]. These differences could be useful to selectively target the different PRDM2 transcripts.…”

“…RNA was reverse transcribed with SuperScript III (Thermofisher) using 500 ng of total RNA. Quantitative RT-PCR analysis was performed as previously reported [ 4 ]. Glyceraldehydes-3-phophate dehydrogenase ( GAPDH ) or peptidylprolyl isomerase A ( PPIA ) were used as housekeeping control genes [ 18 ].…”

“…The pEGFP-C1 vector was purchased from Clontech (Palo Alto, CA, USA) and was used to clone in the BamH1 site, the sequence of RIZ2 open reading frame (NM_015866.4) as reported elsewhere [ 4 , 6 ]. The RIZ2 expressing plasmid, designated pEGFP_hRIZ2, was in frame with the EGFP coding sequence, which was positioned at the N-terminus, without in-frame stop codons.…”

“…Stable clones were selected with 0.8 mg/ml Geneticin- G418 (Sigma-Aldrich). Transfection was verified by fluorescent microscopy and Western blot analyses [ 4 , 6 ].…”

“…Particularly, some PRDMs are endowed with histone methyltransferase activity [1][2][3]. PRDMs are involved in many developmental processes and play key roles in cell differentiation as well as in transducing several signaling pathways [1][2][3][4]. Moreover, numerous studies suggest that the modification of their expression levels, sequence, or structure, has a relevant impact in many human diseases, including cancer [1][2][3].…”