Macrophage differentiation is marked by increased abundance of the mRNA 3’ end processing machinery, altered poly(A) site usage, and sensitivity to the level of CstF64

Abstract

Regulation of mRNA polyadenylation is important for response to external signals and differentiation in several cell types, and results in mRNA isoforms that vary in the amount of coding sequence or 3’ UTR regulatory elements. However, its role in differentiation of monocytes to macrophages has not been investigated. Macrophages are key effectors of the innate immune system that help control infection and promote tissue-repair. However, overactivity of macrophages contributes to pathogenesis of many diseases. In this study, we show that macrophage differentiation is characterized by shortening and lengthening of mRNAs in relevant cellular pathways. The cleavage/polyadenylation (C/P) proteins increase during differentiation, suggesting a possible mechanism for the observed changes in poly(A) site usage. This was surprising since higher C/P protein levels correlate with higher proliferation rates in other systems, but monocytes stop dividing after induction of differentiation. Depletion of CstF64, a C/P protein and known regulator of polyadenylation efficiency, delayed macrophage marker expression, cell cycle exit, attachment, and acquisition of structural complexity, and impeded shortening of mRNAs with functions relevant to macrophage biology. Conversely, CstF64 overexpression increased use of promoter-proximal poly(A) sites and caused the appearance of differentiated phenotypes in the absence of induction. Our findings indicate that regulation of polyadenylation plays an important role in macrophage differentiation.

Elevated pre-mRNA 3′ end processing activity in cancer cells renders vulnerability to inhibition of cleavage and polyadenylation

“…Our comparison of U937 and U937MP cells is particularly revealing about the relative importance of CPA activity vs. cell proliferation for JTE-607-mediated cell death. The quiescent U937MP cells have a higher CPA activity than proliferative U937 cells, which is in line with a recent report showing increased CPA factor expression in macrophage differentiation 50 . As such, U937MP cells display greater transcriptomic distances by JTE-607 than U937 cells.…”

Downregulation of CPSF6 leads to global mRNA 3’ UTR shortening and enhanced antiviral immune responses

“…For example, infection with human cytomegalovirus (HCMV) can induce upregulation of the host polyadenylation factor CPEB1, resulting in shorter 3’ UTRs and longer poly (A) tail lengths of genes in host cells [ 35 ]. Overexpression of CstF64, a 3’ processing factor and known regulator of polyadenylation efficiency, increases the usage of promoter-proximal poly(A) sites and promotes macrophage differentiation without induction [ 36 ]. The 3’ end processing factor NUDT21 limits expression of CD19 in B-cell progenitor acute lymphoblastic leukemia (B-ALL) cells by regulating CD19 mRNA polyadenylation [ 37 ].…”