MicroRNA-155 controls RB phosphorylation in normal and malignant B lymphocytes via the noncanonical TGF-b1/SMAD5 signaling module

MicroRNAs (miRNAs) are important regulators of the human transcriptome, and their role in multiple physiological processes has been well established. Likewise, the participation of these small noncoding RNAs in various pathological states, including cancer, is now fully recognized. Nonetheless, in many instances, the breadth of the dysfunction caused bymiRNA deregulation is not entirely known. In part, this reflects their pleiotropic activities because a singlemiRNA can directly inhibit the expression of dozens, if not hundreds, of genes. However, this may also indicate our limited understanding of the full complement of downstream effects that follow a specific miRNA:target gene interaction. The transforming growth factor b (TGF-b) pathway plays important roles in embryonic development as well as in tissue homeostasis in adult organisms. Thus, deregulation of TGF-b signaling is associated with a variety of human disorders, including cancer. In epithelial malignancies, where the role of TGF-b has been more extensively investigated, the current evidence suggests the presence of an initial tumor suppressive activity, which is followed by an oncogenic profile characterized by epithelial-tomesenchymal transition and metastasis. In contrast, TGF-b signals in normal and malignant B lymphocytes appear to be predominantly suppressive, indicating that deregulation of this pathwaymay contribute to the pathogenesis of B-cell malignancies and interfere with the developmental regulation of normal B cells. We recently described the direct targeting of the transcription factor SMAD5 by microRNA-155 (miR-155). In that report, we showed that miR-155, which is often overexpressed in aggressive B-cell lymphomas, contributes to lymphomagenesis by abrogating the cytostatic effects of the TGF-b pathway toward B lymphocytes. Importantly, although SMAD5 is an integral component of the bone morphogenic protein (BMP) signaling module, we showed that the noncanonical TGF-b1–mediated activation of SMAD5 was present in B-cell lymphomas. Consequently, miR-155 targeting of SMAD5 had broader implications than initially appreciated because it disrupted both BMP and TGF-b signaling. Herein, we examined the downstream effects of the TGF-b1– induced activation of SMAD5 in normal andmalignant B lymphocytes, in the context of gain or loss of miR-155 function. TGF-b1–mediated phosphorylation of SMAD5 led to the transcriptional induction of p21 and p15, a process originally ascribed exclusively to the TGFb1–SMAD2/3 interplay, and decreased the levels of the retinoblastoma protein (RB) phosphorylation. Genetic ablation of SMAD5, p21, or p15 confirmed the relevance of the noncanonical TGFb1–SMAD5 engagement in B lymphocytes. Overexpression of miR-155 in diffuse large B-cell lymphomas (DLBCLs) limited TGF-b1–mediated p21 and p15 induction, resulting in sustained levels of RB hyperphosphorylation and decreased formation of the RB/E2F1 complex. These data suggest that miR-155 can