Biology subjects

Bengoechea, R.

Publications and source records attributed to Bengoechea, R..

4 recordsLinked to original sources

bioRxiv2022

DNAJB6 Isoform Specific Knockdown: Therapeutic Potential for LGMDD1

Dominant missense mutations in DNAJB6, an HSP40 co-chaperone, cause limb girdle muscular dystrophy (LGMD) D1. No treatments are currently available. Two isoforms exist, DNAJB6a and DNAJB6b, each with distinct localizations in muscle. Mutations reside in both isoforms, yet evidence suggests only DNAJB6b is responsible for disease pathogenesis. Mechanistic data supports either a toxic gain of function, a dominant negative mechanism, or a combination of both. Knockdown treatment strategies involving both isoforms carry risk as DNAJB6 knockout is embryonic lethal. We therefore developed an isoform specific knockdown approach using morpholinos. Selective reduction of each isoform was achieved in-vitro in primary mouse myotubes and human myoblasts, as well as in-vivo in mouse skeletal muscle. To assess isoform specific knockdown in LGMDD1, we created primary myotube cultures from a knock-in LGMDD1 mouse model. Using mass spectrometry, we identified an LGMDD1 protein signature related to protein homeostasis and myofibril structure. Selective reduction of DNAJB6b levels in LGMDD1 myotubes corrected much of the proteomic disease signature towards wild type levels. While additional in-vivo functional data is required, these findings suggest selective reduction of DNAJB6b may be a viable therapeutic target for LGMDD1.

neuroscience↗

bioRxiv2022

Loss of function variants in DNAJB4 cause a myopathy with early respiratory failure

DNAJ/HSP40 co-chaperones are integral to the chaperone network, bind client proteins and recruit them to HSP70 for folding. We performed exome sequencing on patients with a presumed hereditary muscle disease and no genetic diagnosis. This identified four individuals from three unrelated families carrying an unreported homozygous stop gain (c.856A>T; p.Lys286Ter), or homozygous missense variants (c.74G>A; p.Arg25Gln and c.785T>C; p.Leu262Ser) in DNAJB4. Affected patients presented with axial rigidity and early respiratory failure requiring ventilator support between the 1st and 4th decade of life. Selective involvement of the semitendinosus and biceps femoris muscles was seen on MRI scans of the thigh. On biopsy, muscle was myopathic with angular fibers, protein inclusions and occasional rimmed vacuoles. DNAJB4 normally localizes to the Z-disc and was absent from muscle and fibroblasts of affected patients supporting a loss of function. Functional studies confirmed that the p.Lys286Ter and p.Leu262Ser mutant proteins are rapidly degraded in cells. In contrast, the p.Arg25Gln mutant protein is stable but failed to complement for DNAJB function in yeast, disaggregate client proteins or protect from heat shock induced cell death consistent with its loss of function. DNAJB4 knockout mice had muscle weakness and fiber atrophy with prominent diaphragm involvement and kyphosis. DNAJB4 knockout muscle and myotubes had myofibrillar disorganization and accumulated Z-disc proteins and protein chaperones. These data demonstrate a novel chaperonopathy associated with DNAJB4 causing a myopathy with early respiratory failure. DNAJB4 loss of function variants may lead to the accumulation of DNAJB4 client proteins resulting in muscle dysfunction and degeneration.

genetics↗

bioRxiv2022

Distinct chaperonopathy in skeletal muscle associated with the dominant variant in DNAJB4

DnaJ homolog, subfamily B, member 4, a member of the heat shock protein 40 chaperones encoded by DNAJB4, is highly expressed in myofibers. We identified a heterozygous c.270 T>A (p.F90L) variant in DNAJB4 in a family with a dominantly inherited distal myopathy, in which affected members have specific features on muscle pathology represented by the presence of cytoplasmic inclusions and the accumulation of desmin, p62, HSP70 and DNAJB4 predominantly in type 1 fibers. Both Dnajb4- F90L knock-in and knockout mice developed muscle weakness and recapitulated the patient muscle pathology in the soleus muscle, where DNAJB4 has the highest expression. These data indicate that the identified variant is causative resulting in defective chaperone function and selective muscle degeneration in specific muscle fibers. This study demonstrates the importance of DNAJB4 in skeletal muscle proteostasis by identifying the associated chaperonopathy.

neuroscience↗

bioRxiv2020

Inhibition of DNAJ-HSP70 interaction improves strength in muscular dystrophy

Dominant mutations in the HSP70 co-chaperone DNAJB6 cause a late onset muscle disease termed limb girdle muscular dystrophy type 1D (LGMD1D), which is characterized by protein aggregation and vacuolar myopathology. Disease mutations reside within the G/F domain of DNAJB6, but the molecular mechanisms underlying dysfunction are not well understood. Using yeast, cell culture, and mouse models of LGMD1D, we find that the toxicity associated with disease-associated DNAJB6 requires its interaction with HSP70, and that abrogating this interaction genetically or with small molecules is protective. In skeletal muscle, DNAJB6 localizes to the Z-disc with HSP70. Whereas HSP70 normally diffuses rapidly between the Z-disc and sarcoplasm, the rate of HSP70s diffusion in LGMD1D mouse muscle is diminished likely because it has an unusual affinity for the Z-disc and mutant DNAJB6. Treating LGMD1D mice with a small molecule inhibitor of the DNAJ-HSP70 complex re-mobilizes HSP70, improves strength and corrects myopathology. These data support a model in which LGMD1D mutations in DNAJB6 are a gain-of-function disease that is, counter-intuitively, mediated via HSP70 binding. Thus, therapeutic approaches targeting HSP70:DNAJB6 may be effective in treating this inherited muscular dystrophy.

cell biology↗