Transgenic expression of Hsc70 in pancreatic islets enhances autoimmune diabetes in response to beta cell damage.

Inflammation following tissue damage promotes lymphocyte recruitment, tissue remodeling, and wound healing while maintaining self tolerance. Endogenous signals associated with tissue damage and cell death have been proposed to initiate and instruct immune responses following injury. In this study, we have examined the effects of elevated levels of a candidate endogenous danger signal, heat shock cognate protein 70 (hsc70), on stimulation of inflammation and autoimmunity following cell damage. We find that damage to pancreatic beta cells expressing additional cytosolic hsc70 leads to an increased incidence of diabetes in a transgenic mouse model. Steady-state levels of activated APC and T cell populations in the draining lymph node were enhanced, which further increased following streptozotocin-induced beta cell death. In addition, proinflammatory serum cytokines, and lymphocyte recruitment were increased in hsc70 transgenic mice. Islet Ag-specific T cells underwent a greater extent of proliferation in the lymph nodes of mice expressing hsc70 following beta cell damage, suggesting elevated Ag presentation following release of Ag in the presence of hsc70. These findings suggest that an elevated content of hsc70 in cells undergoing necrotic or apoptotic cell death can increase the extent of sterile inflammation and increase the susceptibility to autoimmunity.

Immunomodulatory drugs disrupt the cereblon-CD147-MCT1 axis to exert antitumor activity and teratogenicity.

Immunomodulatory drugs (IMiDs), such as thalidomide and its derivatives lenalidomide and pomalidomide, are key treatment modalities for hematologic malignancies, particularly multiple myeloma (MM) and del(5q) myelodysplastic syndrome (MDS). Cereblon (CRBN), a substrate receptor of the CRL4 ubiquitin ligase complex, is the primary target by which IMiDs mediate anticancer and teratogenic effects. Here we identify a ubiquitin-independent physiological chaperone-like function of CRBN that promotes maturation of the basigin (BSG; also known as CD147) and solute carrier family 16 member 1 (SLC16A1; also known as MCT1) proteins. This process allows for the formation and activation of the CD147-MCT1 transmembrane complex, which promotes various biological functions, including angiogenesis, proliferation, invasion and lactate export. We found that IMiDs outcompete CRBN for binding to CD147 and MCT1, leading to destabilization of the CD147-MCT1 complex. Accordingly, IMiD-sensitive MM cells lose CD147 and MCT1 expression after being exposed to IMiDs, whereas IMiD-resistant cells retain their expression. Furthermore, del(5q) MDS cells have elevated CD147 expression, which is attenuated after IMiD treatment. Finally, we show that BSG (CD147) knockdown phenocopies the teratogenic effects of thalidomide exposure in zebrafish. These findings provide a common mechanistic framework to explain both the teratogenic and pleiotropic antitumor effects of IMiDs.

Disruption of the PRKCD-FBXO25-HAX-1 axis attenuates the apoptotic response and drives lymphomagenesis.

We searched for genetic alterations in human B cell lymphoma that affect the ubiquitin-proteasome system. This approach identified FBXO25 within a minimal common region of frequent deletion in mantle cell lymphoma (MCL). FBXO25 encodes an orphan F-box protein that determines the substrate specificity of the SCF (SKP1-CUL1-F-box)(FBXO25) ubiquitin ligase complex. An unbiased screen uncovered the prosurvival protein HCLS1-associated protein X-1 (HAX-1) as the bona fide substrate of FBXO25 that is targeted after apoptotic stresses. Protein kinase Cδ (PRKCD) initiates this process by phosphorylating FBXO25 and HAX-1, thereby spatially directing nuclear FBXO25 to mitochondrial HAX-1. Our analyses in primary human MCL identify monoallelic loss of FBXO25 and stabilizing HAX1 phosphodegron mutations. Accordingly, FBXO25 re-expression in FBXO25-deleted MCL cells promotes cell death, whereas expression of the HAX-1 phosphodegron mutant inhibits apoptosis. In addition, knockdown of FBXO25 significantly accelerated lymphoma development in Eµ-Myc mice and in a human MCL xenotransplant model. Together we identify a PRKCD-dependent proapoptotic mechanism controlling HAX-1 stability, and we propose that FBXO25 functions as a haploinsufficient tumor suppressor and that HAX1 is a proto-oncogene in MCL.

SCFFbxo9 and CK2 direct the cellular response to growth factor withdrawal via Tel2/Tti1 degradation and promote survival in multiple myeloma.

The Tel2 (also known as Telo2) and Tti1 proteins control the cellular abundance of mammalian PIKKs and are integral components of mTORC1 and mTORC2. Here we report that Tel2 and Tti1 are targeted for degradation within mTORC1 by the SCFFbxo9 ubiquitin ligase to adjust mTOR signalling to growth factor availability. This process is primed by CK2, which translocates to the cytoplasm to mediate mTORC1-specific phosphorylation of Tel2/Tti1, subsequent to growth factor deprivation. As a consequence, mTORC1 is inactivated to restrain cell growth and protein translation whereas relief of feedback inhibition activates the PI(3)K/TORC2/Akt pathway to sustain survival. Significantly, primary human multiple myelomas exhibit high levels of Fbxo9. In this setting, PI(3)K/TORC2/Akt signalling and survival of multiple myeloma cells is dependent on Fbxo9 expression. Thus, mTORC1-specific degradation of the Tel2 and Tti1 proteins represents a central mTOR regulatory mechanism with implications in multiple myeloma, both in promoting survival and in providing targets for the specific treatment of multiple myeloma with high levels of Fbxo9 expression.

Frequency encoding of cholinergic- and purinergic-mediated signaling to mouse urinary bladder smooth muscle: modulation by BK channels.

In the urinary bladder, contractions of the detrusor muscle and urine voiding are induced by the neurotransmitters ACh and ATP, released from parasympathetic nerves. Activation of K(+) channels, in particular the large-conductance Ca(2+)-activated K(+) (BK) channels, opposes increases in excitability and contractility of urinary bladder smooth muscle (UBSM). We have shown that deleting the gene mSlo1 in mice (Slo(-/-)), encoding the BK channel, leads to enhanced nerve-mediated and neurotransmitter-dependent contractility of UBSM (38). Here, we examine the location of the BK channel in urinary bladder strips from mouse. Immunohistochemical analysis revealed that the channel is expressed in UBSM but not in nerves that innervate the smooth muscle. The relationship between electrical field stimulation and force generation of the cholinergic and purinergic pathways was examined by applying blockers of the respective receptors in UBSM strips from wild-type and from Slo(-/-) (knockout) mice. In wild-type strips, the stimulation frequency required to obtain a half-maximal force was significantly lower for the purinergic (7.2 +/- 0.3 Hz) than the cholinergic pathway (19.1 +/- 1.5 Hz), whereas the maximum force was similar. Blocking BK channels with iberiotoxin or ablation of the Slo gene increased cholinergic- and purinergic-mediated force at low frequencies, i.e., significantly decreased the frequency for a half-maximal force. Our results indicate that the BK channel has a very significant role in reducing both cholinergic- and purinergic-induced contractility and suggest that alterations in BK channel expression or function could contribute to pathologies such as overactive detrusor.