Molecular basis for Nse5-6 mediated regulation of Smc5/6 functions.

The Smc5/6 complex (Smc5/6) is important for genome replication and repair in eukaryotes. Its cellular functions are closely linked to the ATPase activity of the Smc5 and Smc6 subunits. This activity requires the dimerization of the motor domains of the two SMC subunits and is regulated by the six non-SMC subunits (Nse1 to Nse6). Among the NSEs, Nse5 and Nse6 form a stable subcomplex (Nse5-6) that dampens the ATPase activity of the complex. However, the underlying mechanisms and biological significance of this regulation remain unclear. Here, we address these issues using structural and functional studies. We determined cryo-EM structures of the yeast Smc5/6 derived from complexes consisting of either all eight subunits or a subset of five subunits. Both structures reveal that Nse5-6 associates with Smc6's motor domain and the adjacent coiled-coil segment, termed the neck region. Our structural analyses reveal that this binding is compatible with motor domain dimerization but results in dislodging the Nse4 subunit from the Smc6 neck. As the Nse4-Smc6 neck interaction favors motor domain engagement and thus ATPase activity, Nse6's competition with Nse4 can explain how Nse5-6 disfavors ATPase activity. Such regulation could in principle differentially affect Smc5/6-mediated processes depending on their needs of the complex's ATPase activity. Indeed, mutagenesis data in cells provide evidence that the Nse6-Smc6 neck interaction is important for the resolution of DNA repair intermediates but not for replication termination. Our results thus provide a molecular basis for how Nse5-6 modulates the ATPase activity and cellular functions of Smc5/6.

MCOLN1 gene therapy corrects neurologic dysfunction in the mouse model of mucolipidosis IV.

Mucolipidosis IV (MLIV) is an orphan disease leading to debilitating psychomotor deficits and vision loss. It is caused by loss-of-function mutations in the MCOLN1 gene that encodes the lysosomal transient receptor potential channel mucolipin1, or TRPML1. With no existing therapy, the unmet need in this disease is very high. Here, we showed that AAV-mediated CNS-targeted gene transfer of the human MCOLN1 gene rescued motor function and alleviated brain pathology in the MLIV mouse model. Using the AAV-PHP.b vector in symptomatic mice, we showed long-term reversal of declined motor function and significant delay of paralysis. Next, using self-complementary AAV9 clinical candidate vector, we showed that its intracerebroventricular administration in post-natal day 1 mice significantly improved motor function, myelination and reduced lysosomal storage load in the MLIV mouse brain. Based on our data and general advancements in the gene therapy field, we propose scAAV9-mediated CSF-targeted MCOLN1 gene transfer as a therapeutic strategy in MLIV.

Patient-Reported Outcomes Are Better after Oncoplastic Breast Conservation than after Mastectomy and Autologous Reconstruction.

Oncoplastic breast-conserving surgery (OBCS) avoids mastectomy for larger tumors, but patient-reported outcomes are unknown. METHODS: The BREAST-Q questionnaire was distributed to 333 women following therapeutic mammaplasty or latissimus dorsi (LD) miniflap since 1991 [tumor diameter, 32.5 (5-100) mm). QScore software generated scores/100 for breast appearance, physical, emotional, and sexual wellbeing. Outcomes following therapeutic mammaplasty and LD miniflap were compared and qualitative data analyzed to identify common themes relating to satisfaction. RESULTS: One hundred fifty (45%) women responded [mammaplasty versus LD miniflap, 52% versus 42%; age, 52 (30-83) years; follow-up, 84 (4-281) months). Eighty-nine percent rated OBCS better than mastectomy, > 80% recommending it to others. Mean outcome scores for breast appearance, physical, and emotional wellbeing were high and persisted beyond 15 years. Therapeutic mammaplasty patients were significantly more satisfied than those undergoing LD miniflap with the shape (P < 0.05), the size (P < 0.005), and the natural feel of the treated breast (P = 0.01). They demonstrated similar scores for physical and emotional wellbeing and a lower score for sexual wellbeing than LD miniflap patients. More LD miniflap patients reported back/shoulder symptoms and were more likely to report upper back pain (P < 0.05), but very few (< 5%) were concerned about donor-site appearance. Overall satisfaction with surgical outcomes was high in both OBCS groups (82% "excellent/very good") but greatest after therapeutic mammaplasty (P < 0.005). CONCLUSIONS: Patients report long-lasting satisfaction after OBCS and outcomes that compare very favorably with those reported following mastectomy and immediate autologous reconstruction.