Heat-shock protein (Hsp) 90 assists in the folding of diverse sets of client proteins including kinases and growth hormone receptors. Hsp70 plays a major role in many Hsp90 functions by interacting and modulating conformation of its substrates before being transferred to Hsp90s for final maturation. Each eukaryote contains multiple members of the Hsp70 family. However, the role of different Hsp70 isoforms in Hsp90 chaperoning actions remains unknown. Using v-Src as an Hsp90 substrate, we examined the role of each of the four yeast cytosolic Ssa Hsp70s in regulating Hsp90 functions. We show that the strain expressing stress-inducible Ssa3 or Ssa4, and the not constitutively expressed Ssa1 or Ssa2, as the sole Ssa Hsp70 isoform reduces v-Src-mediated growth defects. The study shows that although different Hsp70 isoforms interact similarly with Hsp90s, v-Src maturation is less efficient in strains expressing Ssa4 as the sole Hsp70. We further show that the functional distinction between Ssa2 and Ssa4 is regulated by its C-terminal domain. Further studies reveal that Ydj1, which is known to assist substrate transfer to Hsp70s, interacts relatively weakly with Ssa4 compared with Ssa2, which could be the basis for poor maturation of the Hsp90 client in cells expressing stress-inducible Ssa4 as the sole Ssa Hsp70. The study thus reveals a novel role of Ydj1 in determining the functional distinction among Hsp70 isoforms with respect to the Hsp90 chaperoning action.

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Haemorrhagic stroke represents a significant public health burden, yet our knowledge and ability to treat this type of stroke are lacking. Previously we showed that we can target ischaemic-stroke lesions by selective translocation of lipid nanoparticles through the site of blood-brain barrier (BBB) disruption. The data we presented in this study provide compelling evidence that haemorrhagic stroke in mice induces BBB injury that mimics key features of the human pathology and, more importantly, provides a gate for entry of lipid nanoparticles-based therapeutics selectively to the bleeding site. Methods: Haemorrhagic stroke was induced in mice by intra-striatal collagenase injection. lipid nanoparticles were injected intravenously at 3 h, 24 h & 48 h post-haemorrhagic stroke and accumulation in the brain studied using in-vivo optical imaging and histology. BBB integrity, brain water content and iron accumulation were characterised using dynamic contrast-enhanced MRI, quantitative T 1 mapping, and gradient echo MRI. Results: Using in-vivo SPECT/CT imaging and optical imaging revealed biphasic lipid nanoparticles entry into the bleeding site, with an early phase of increased uptake at 3-24 h post-haemorrhagic stroke, followed by a second phase at 48-72 h. Lipid nanoparticles entry into the brain post-haemorrhage showed an identical entry pattern to the trans-BBB leakage rate (K trans [min -1 ]) of Gd-DOTA, a biomarker for BBB disruption, measured using dynamic contrast-enhanced MRI. Discussion: Our findings suggest that selective accumulation of liposomes into the lesion site is linked to a biphasic pattern of BBB hyper-permeability. This approach provides a unique opportunity to selectively and efficiently deliver therapeutic molecules across the BBB, an approach that has not been utilised for haemorrhagic stroke therapy and is not achievable using free small drug molecules.

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