Transforming growth factor-␤ (TGF-␤

The mammalian target of rapamycin (mTOR) pathway is commonly activated in human cancers. The activity of mTOR complex 1 (mTORC1) signaling is supported by the intracellular positioning of cellular compartments and vesicle trafficking, regulated by Rab GTPases. Here we showed that tuftelin 1 (TUFT1) was involved in the activation of mTORC1 through modulating the Rab GTPase-regulated process. TUFT1 promoted tumor growth and metastasis. Consistently, the expression of TUFT1 correlated with poor prognosis in lung, breast and gastric cancers. Mechanistically, TUFT1 physically interacted with RABGAP1, thereby modulating intracellular lysosomal positioning and vesicular trafficking, and promoted mTORC1 signaling. In addition, expression of TUFT1 predicted sensitivity to perifosine, an alkylphospholipid that alters the composition of lipid rafts. Perifosine treatment altered the positioning and trafficking of cellular compartments to inhibit mTORC1. Our observations indicate that TUFT1 is a key regulator of the mTORC1 pathway and suggest that it is a promising therapeutic target or a biomarker for tumor progression.

p53 functions to induce cellular senescence, which is incompatible with self-renewal of pluripotent stem cells such as induced pluripotent stem cells (iPSC) and embryonic stem cells (ESC). However, p53 also has essential roles in these cells through DNA damage repair for maintaining genomic integrity and high sensitivity to apoptosis for eliminating severely damaged cells. We hypothesized that Δ133p53, a physiological inhibitory p53 isoform, is involved in the balanced regulation of self-renewing capacity, DNA damage repair and apoptosis. We examined 12 lines of human iPSC and their original fibroblasts, as well as three ESC lines, for endogenous protein levels of Δ133p53 and full-length p53 (FL-p53), and mRNA levels of various p53 target genes. While FL-p53 levels in iPSC and ESC widely ranged from below to above those in the fibroblasts, all iPSC and ESC lines expressed elevated levels of Δ133p53. The p53-inducible genes that mediate cellular senescence (p21, miR-34a, PAI-1 and IGFBP7), but not those for apoptosis (BAX and PUMA) and DNA damage repair (p53R2), were downregulated in iPSC and ESC. Consistent with these endogenous expression profiles, overexpression of Δ133p53 in human fibroblasts preferentially repressed the p53-inducible senescence mediators and significantly enhanced their reprogramming to iPSC. The iPSC lines derived from Δ133p53-overexpressing fibroblasts formed well-differentiated, benign teratomas in immunodeficient mice and had fewer numbers of somatic mutations than an iPSC derived from p53-knocked-down fibroblasts, suggesting that Δ133p53 overexpression is non- or less oncogenic and mutagenic than total inhibition of p53 activities. Overexpressed Δ133p53 prevented FL-p53 from binding to the regulatory regions of p21 and miR-34a promoters, providing a mechanistic basis for its dominant-negative inhibition of a subset of p53 target genes. This study supports the hypothesis that upregulation of Δ133p53 is an endogenous mechanism that facilitates human somatic cells to become self-renewing pluripotent stem cells with maintained apoptotic and DNA repair activities.

Cellular senescence is a hallmark of normal aging and aging-related syndromes, including the premature aging disorder Hutchinson-Gilford Progeria Syndrome (HGPS), a rare genetic disorder caused by a single mutation in the LMNA gene that results in the constitutive expression of a truncated splicing mutant of lamin A known as progerin. Progerin accumulation leads to increased cellular stresses including unrepaired DNA damage, activation of the p53 signaling pathway and accelerated senescence. We previously established that the p53 isoforms Δ133p53 and p53β regulate senescence in normal human cells. However, their role in premature aging is unknown. Here, we report that p53 isoforms are expressed in primary fibroblasts derived from HGPS patients, are associated with their accelerated senescence and that their manipulation can restore the replication capacity of HGPS fibroblasts. We found that in near-senescent HGPS fibroblasts, which exhibit low levels of Δ133p53 and high levels of p53β, restoration of Δ133p53 expression was sufficient to extend replicative lifespan and delay senescence, despite progerin levels and abnormal nuclear morphology remaining unchanged. Conversely, Δ133p53 depletion or p53β overexpression accelerated the onset of senescence in otherwise proliferative HGPS fibroblasts. Our data indicate that Δ133p53 exerts its role by modulating full-length p53 (FLp53) signaling to extend the replicative lifespan and promotes the repair of spontaneous progerin-induced DNA double strand breaks (DSBs). We showed that Δ133p53 dominant-negative inhibition of FLp53 occurs directly at the p21/CDKN1A and miR-34a promoters, two p53-senescence associated genes. In addition, Δ133p53 expression increased expression of the DNA repair RAD51, likely through upregulation of E2F1, a transcription factor that activates RAD51, to promote repair of DSBs. In summary, our data indicate that Δ133p53 modulates p53 signaling to repress progerin-induced early onset of senescence in HGPS cells. Therefore, restoration of Δ133p53 expression may be a novel therapeutic strategy to treat aging-associated phenotypes of HGPS in vivo.

Transforming growth factor (TGF)‐β induces various cellular responses principally through Smad‐dependent transcriptional regulation. Activated Smad complexes cooperate with transcription factors in regulating a group of target genes. The target genes controlled by the same Smad‐cofactor complexes are denoted a synexpression group. We found that an Id‐like helix‐loop‐helix protein, human homologue of Maid (HHM), is a synexpression group‐restricted regulator of TGF‐β signalling. HHM suppressed TGF‐β‐induced growth inhibition and cell migration but not epithelial—mesenchymal transition. In addition, HHM inhibited TGF‐β‐induced expression of plasminogen activator inhibitor‐type 1 (PAI‐1), PDGF‐B, and p21WAF, but not Snail. We identified a basic‐helix‐loop‐helix protein, Olig1, as one of the Smad‐binding transcription factors affected by HHM. Olig1 interacted with Smad2/3 in response to TGF‐β stimulation, and was involved in transcriptional activation of PAI‐1 and PDGF‐B. HHM, but not Id proteins, inhibited TGF‐β signalling‐dependent association of Olig1 with Smad2/3 through physical interaction with Olig1. HHM thus appears to regulate a subset of TGF‐β target genes including the Olig1‐Smad synexpression group. HHM is the first example of a cellular response‐selective regulator of TGF‐β signalling with clearly determined mechanisms.