Caspase-independent cell death: an anti-cancer double-whammy

No abstract available

Ubiquitination and proteasomal degradation of ATG12 regulates its proapoptotic activity

During macroautophagy, conjugation of ATG12 to ATG5 is essential for LC3 lipidation and autophagosome formation. Additionally, ATG12 has ATG5-independent functions in diverse processes including mitochondrial fusion and mitochondrial-dependent apoptosis. In this study, we investigated the regulation of free ATG12. In stark contrast to the stable ATG12–ATG5 conjugate, we find that free ATG12 is highly unstable and rapidly degraded in a proteasome-dependent manner. Surprisingly, ATG12, itself a ubiquitin-like protein, is directly ubiquitinated and this promotes its proteasomal degradation. As a functional consequence of its turnover, accumulation of free ATG12 contributes to proteasome inhibitor-mediated apoptosis, a finding that may be clinically important given the use of proteasome inhibitors as anticancer agents. Collectively, our results reveal a novel interconnection between autophagy, proteasome activity, and cell death mediated by the ubiquitin-like properties of ATG12

Mitochondrial permeabilization engages NF-κB-dependent anti-tumour activity under caspase deficiency

Apoptosis represents a key anti-cancer therapeutic effector mechanism. During apoptosis, mitochondrial outer membrane permeabilization (MOMP) typically kills cells even in the absence of caspase activity. Caspase activity can also have a variety of unwanted consequences that include DNA damage. We therefore investigated whether MOMP-induced caspase-independent cell death (CICD) might be a better way to kill cancer cells. We find that cells undergoing CICD display potent pro-inflammatory effects relative to apoptosis. Underlying this, MOMP was found to stimulate NF-κB activity through the downregulation of inhibitor of apoptosis proteins. Strikingly, engagement of CICD displays potent anti-tumorigenic effects, often promoting complete tumour regression in a manner dependent on intact immunity. Our data demonstrate that by activating NF-κB, MOMP can exert additional signalling functions besides triggering cell death. Moreover, they support a rationale for engaging caspase-independent cell death in cell-killing anti-cancer therapies

The HUSH complex is a gatekeeper of type I interferon through epigenetic regulation of LINE-1s

Funder: UKRI Future Leaders Fellowship (MR/S034498/1)Abstract: The Human Silencing Hub (HUSH) complex is necessary for epigenetic repression of LINE-1 elements. We show that HUSH-depletion in human cell lines and primary fibroblasts leads to induction of interferon-stimulated genes (ISGs) through JAK/STAT signaling. This effect is mainly attributed to MDA5 and RIG-I sensing of double-stranded RNAs (dsRNAs). This coincides with upregulation of primate-conserved LINE-1s, as well as increased expression of full-length hominid-specific LINE-1s that produce bidirectional RNAs, which may form dsRNA. Notably, LTRs nearby ISGs are derepressed likely rendering these genes more responsive to interferon. LINE-1 shRNAs can abrogate the HUSH-dependent response, while overexpression of an engineered LINE-1 construct activates interferon signaling. Finally, we show that the HUSH component, MPP8 is frequently downregulated in diverse cancers and that its depletion leads to DNA damage. These results suggest that LINE-1s may drive physiological or autoinflammatory responses through dsRNA sensing and gene-regulatory roles and are controlled by the HUSH complex

Coordination by Cdc42 of actin, contractility, and adhesion for melanoblast movement in mouse skin

YesThe individual molecular pathways downstream of Cdc42, Rac, and Rho GTPases are well documented, but we know surprisingly little about how these pathways are coordinated when cells move in a complex environment in vivo. In the developing embryo, melanoblasts originating from the neural crest must traverse the dermis to reach the epidermis of the skin and hair follicles. We previously established that Rac1 signals via Scar/WAVE and Arp2/3 to effect pseudopod extension and migration of melanoblasts in skin. Here we show that RhoA is redundant in the melanocyte lineage but that Cdc42 coordinates multiple motility systems independent of Rac1. Similar to Rac1 knockouts, Cdc42 null mice displayed a severe loss of pigmentation, and melanoblasts showed cell-cycle progression, migration, and cytokinesis defects. However, unlike Rac1 knockouts, Cdc42 null melanoblasts were elongated and displayed large, bulky pseudopods with dynamic actin bursts. Despite assuming an elongated shape usually associated with fast mesenchymal motility, Cdc42 knockout melanoblasts migrated slowly and inefficiently in the epidermis, with nearly static pseudopods. Although much of the basic actin machinery was intact, Cdc42 null cells lacked the ability to polarize their Golgi and coordinate motility systems for efficient movement. Loss of Cdc42 de-coupled three main systems: actin assembly via the formin FMNL2 and Arp2/3, active myosin-II localization, and integrin-based adhesion dynamics.Cancer Research UK (to L.M.M. [A17196], R.H.I. [A19257], and S.W.G.T.) and NIH grants P01-GM103723 and P41-EB002025 (to K.M.H.). N.R.P. is supported by a Pancreatic Cancer Research Fund grant (to L.M.M.). Funding to Prof. Rottner by the Deutsche Forschungsgemeinschaft (grant RO2414/3-2)

A macrophage-pericyte axis directs tissue restoration via amphiregulin-induced transforming growth factor beta activation

The epidermal growth factor receptor ligand Amphiregulin has a well-documented role in the restoration of tissue homeostasis after injury; however, the mechanism by which Amphiregulin contributes to wound repair remains unknown. Here we show that Amphiregulin functioned by releasing bioactive transforming growth factor beta (TGF-β) from latent complexes via integrin-αV activation. Using acute injury models in two different tissues, we found that by inducing TGF-β activation on mesenchymal stromal cells (pericytes), Amphiregulin induced their differentiation into myofibroblasts, thereby selectively contributing to the restoration of vascular barrier function within injured tissue. Furthermore, we identified macrophages as a critical source of Amphiregulin, revealing a direct effector mechanism by which these cells contribute to tissue restoration after acute injury. Combined, these observations expose a so far under-appreciated mechanism of how cells of the immune system selectively control the differentiation of tissue progenitor cells during tissue repair and inflammation

Investigating non-apoptotic cell death in cancer

No abstract available

Mito-priming as a method of engineered Bcl-2 addiction

Most apoptotic stimuli require mitochondrial outer membrane permeabilization (MOMP) in order to execute cell death. As such, MOMP is subject to tight control by Bcl-2 family proteins. We have developed a powerful new technique to investigate Bcl-2-mediated regulation of MOMP. This method, called mito-priming, uses co-expression of pro- and anti-apoptotic Bcl-2 proteins to engineer Bcl-2 addiction. On addition of Bcl-2 targeting BH3 mimetics, mito-primed cells undergo apoptosis in a rapid and synchronous manner. Using this method we have comprehensively surveyed the efficacy of BH3 mimetic compounds, identifying potent and specific MCL-1 inhibitors. Furthermore, by combining different pro- and anti-apoptotic Bcl-2 pairings together with CRISPR/Cas9-based genome editing, we find that tBID and PUMA can preferentially kill in a BAK-dependent manner. In summary, mito-priming represents a facile and robust means to trigger mitochondrial apoptosis