A new baby in the c-Myc-directed transcriptional machinery: Che-1/AATF

B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most common malignancy in childhood. Despite the high cure-rate, identifying new druggable molecular targets is still of great interest. In a cohort of BCP-ALL pediatric patients, irrespectively of the molecule/karyotype lesions found, we recently observed high expression of c-Myc and Che-1/AATF, which disappears at time of remission. Study of the molecular mechanisms involved in this co-expression revealed that Che-1 expression was crucial for induction of blast-cell proliferation driven by c-Myc. Furthermore, Che-1/AATF silencing in primary BCP-ALL cell lines improves responsiveness to chemotherapy. These data individuate Che-1 as a possible novel target in the treatment of BCP-ALL able to affect c-Myc-driven tumorigenicity.     

Abnormal regulation of TSG101 in mice with spongiform neurodegeneration

Spongiform neurodegeneration is characterized by the appearance of vacuoles throughout the central nervous system. It has many potential causes, but the underlying cellular mechanisms are not well understood. Mice lacking the E3 ubiquitin ligase Mahogunin Ring Finger-1 (MGRN1) develop age-dependent spongiform encephalopathy. We identified an interaction between a “PSAP” motif in MGRN1 and the ubiquitin E2 variant (UEV) domain of TSG101, a component of the endosomal sorting complex required for transport I (ESCRT-I), and demonstrate that MGRN1 multimonoubiquitinates TSG101. We examined the in vivo consequences of loss of MGRN1 on TSG101 expression and function in the mouse brain. The pattern of TSG101 ubiquitination differed in the brains of wild-type mice and Mgrn1 null mutant mice: at 1 month of age, null mutant mice had less ubiquitinated TSG101, while in adults, mutant mice had more ubiquitinated, insoluble TSG101 than wild-type mice. There was an associated increase in epidermal growth factor receptor (EGFR) levels in mutant brains. These results suggest that loss of MGRN1 promotes ubiquitination of TSG101 by other E3s and may prevent its disassociation from endosomal membranes or cause it to form insoluble aggregates. Our data implicate loss of normal TSG101 function in endo-lysosomal trafficking in the pathogenesis of spongiform neurodegeneration in Mgrn1 null mutant mice.     

拮抗凋亡转录因子与肿瘤增殖及凋亡

肿瘤是目前临床常见的疾病之一.肿瘤的经典治疗方式主要包括手术切除、放疗和化疗.近年来肿瘤治疗的手段不断发展,但许多进展期的肿瘤仍然未见较有效的治疗方式,因此亟需新的治疗手段.肿瘤细胞的特点是具有无限增殖和抵抗凋亡的能力.因此,鉴定参与肿瘤细胞增殖和凋亡的基因将为肿瘤治疗提供潜在的靶点.拮抗凋亡转录因子(apoptosi...     

An HRD/DER-independent ER quality control mechanism involves Rsp5p-dependent ubiquitination and ER-Golgi transport

There is increasing evidence that ubiquitination of receptors provides an important endosomal sorting signal. Here we report that mammalian class E vacuolar protein-sorting (vps) proteins recognize ubiquitin. Both tumor susceptibility gene 101 (TSG101)/human VPS (hVPS)28 and hepatocyte growth factor receptor substrate (Hrs) cytosolic complexes bind ubiquitin-agarose. TSG101 and hVPS28 are localized to endosomes that contain internalized EGF receptor and label strongly for ubiquitinated proteins. Microinjection of anti-hVPS28 specifically retards EGF degradation and leads to endosomal accumulation of ubiquitin-protein conjugates. Likewise, depletion of TSG101 impairs EGF trafficking and causes dramatic relocalization of ubiquitin to endocytic compartments. Similar defects are found in cells overexpressing Hrs, further emphasizing the links between class E protein function, receptor trafficking, and endosomal ubiquitination.     

Tumor susceptibility gene 101 (TSG101) is a novel binding-partner for the class II Rab11-FIPs

The Rab11-FIPs (Rab11-family interacting proteins; henceforth, FIPs) are a family of Rab11a/Rab11b/Rab25 GTPase effector proteins implicated in an assortment of intracellular trafficking processes. Through proteomic screening, we have identified TSG101 (tumor susceptibility gene 101), a component of the ESCRT-I (endosomal sorting complex required for transport) complex, as a novel FIP4-binding protein, which we find can also bind FIP3. We show that α-helical coiled-coil regions of both TSG101 and FIP4 mediate the interaction with the cognate protein, and that point mutations in the coiled-coil regions of both TSG101 and FIP4 abrogate the interaction. We find that expression of TSG101 and FIP4 mutants cause cytokinesis defects, but that the TSG101-FIP4 interaction is not required for localisation of TSG101 to the midbody/Flemming body during abscission. Together, these data suggest functional overlap between Rab11-controlled processes and components of the ESCRT pathway.     

Lysosomal localization of ubiquitinated Jun requires multiple determinants in a lysine-27-linked polyubiquitin conjugate

Ubiquitination regulates many cellular functions, including protein localization and degradation. Each function is specified by unique determinants in the conjugate. Ubiquitinated Jun is localized to lysosomes for degradation. Here, we characterized determinants of Jun ubiquitination and lysosomal localization by using ubiquitin-mediated fluorescence complementation (UbFC) in living cells and analysis of the stoichiometry of ubiquitin linked to Jun extracted from cells. The δ region of Jun and isoleucine-44 in ubiquitin were required for lysosomal localization of the conjugate. Ubiquitin containing only lysine-27, but no other single-lysine ubiquitin, mediated Jun ubiquitination, albeit at lower stoichiometry than wild-type ubiquitin. These conjugates were predominantly nuclear, but coexpression of lysine-27 and lysine-less ubiquitins enhanced the mean stoichiometry of Jun ubiquitination and lysosomal localization of the conjugate. Hepatocyte growth factor-regulated tyrosine kinase substrate (HRS) and tumor susceptibility gene 101 (TSG101) colocalized with ubiquitinated Jun. Knockdown of HRS or TSG101 inhibited lysosomal localization of ubiquitinated Jun and reduced Jun turnover. Ubiquitination of other Fos and Jun family proteins had distinct effects on their localization. Our results indicate that Jun is polyubiquitinated by E3 ligases that produce lysine-27–linked chains. Lysosomal localization of the conjugate requires determinants in Jun and in ubiquitin that are recognized in part by TSG101 and HRS, facilitating selective translocation and degradation of ubiquitinated Jun.