Balanced activities of Hsp70 and the ubiquitin proteasome system underlie cellular protein homeostasis.

To counteract proteotoxic stress and cellular aging, protein quality control (PQC) systems rely on the refolding, degradation and sequestration of misfolded proteins. In the Hsp70 chaperone system plays a central role in protein refolding, while degradation is predominantly executed by the ubiquitin proteasome system (UPS). The sequestrases Hsp42 and Btn2 deposit misfolded proteins in cytosolic and nuclear inclusions, thereby restricting the accessibility of misfolded proteins to Hsp70 and preventing the exhaustion of limited Hsp70 resources.

Cellular sequestrases maintain basal Hsp70 capacity ensuring balanced proteostasis.

Maintenance of cellular proteostasis is achieved by a multi-layered quality control network, which counteracts the accumulation of misfolded proteins by refolding and degradation pathways. The organized sequestration of misfolded proteins, actively promoted by cellular sequestrases, represents a third strategy of quality control. Here we determine the role of sequestration within the proteostasis network in Saccharomyces cerevisiae and the mechanism by which it occurs.

A prion-like domain in Hsp42 drives chaperone-facilitated aggregation of misfolded proteins.

Chaperones with aggregase activity promote and organize the aggregation of misfolded proteins and their deposition at specific intracellular sites. This activity represents a novel cytoprotective strategy of protein quality control systems; however, little is known about its mechanism. In yeast, the small heat shock protein Hsp42 orchestrates the stress-induced sequestration of misfolded proteins into cytosolic aggregates (CytoQ).

Small heat shock proteins sequester misfolding proteins in near-native conformation for cellular protection and efficient refolding.

Small heat shock proteins (sHsp) constitute an evolutionary conserved yet diverse family of chaperones acting as first line of defence against proteotoxic stress. sHsps coaggregate with misfolded proteins but the molecular basis and functional implications of these interactions, as well as potential sHsp specific differences, are poorly explored. In a comparative analysis of the two yeast sHsps, Hsp26 and Hsp42, we show in vitro that model substrates retain near-native state and are kept physically separated when complexed with either sHsp, while being completely unfolded when aggregated without sHsps.

Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers.

Tandem fluorescent protein timers (tFTs) report on protein age through time-dependent change in color, which can be exploited to study protein turnover and trafficking. Each tFT, composed of two fluorescent proteins (FPs) that differ in maturation kinetics, is suited to follow protein dynamics within a specific time range determined by the maturation rates of both FPs. So far, tFTs have been constructed by combining slower-maturing red fluorescent proteins (redFPs) with the faster-maturing superfolder green fluorescent protein (sfGFP).

Compartment-specific aggregases direct distinct nuclear and cytoplasmic aggregate deposition.

Disruption of the functional protein balance in living cells activates protective quality control systems to repair damaged proteins or sequester potentially cytotoxic misfolded proteins into aggregates. The established model based on Saccharomyces cerevisiae indicates that aggregating proteins in the cytosol of eukaryotic cells partition between cytosolic juxtanuclear (JUNQ) and peripheral deposits. Substrate ubiquitination acts as the sorting principle determining JUNQ deposition and subsequent degradation.

Fabrication of optical filters based on polymer asymmetric Bragg couplers.

In this work, we successfully developed a process to fabricate dual-channel polymeric waveguide filters based on an asymmetric Bragg coupler (ABC) using holographic interference techniques, soft lithography, and micro molding. At the cross- and self-reflection Bragg wavelengths, the transmission dips of approximately -16.4 and -11.

Fabrication of high-resolution periodical structure on polymer waveguides using a replication process.

This paper describes a procedure to replicate a polymeric wavelength filter. In this work, the grating structure on a polymer is fabricated first using holographic interferometry and micro-molding processes. The polymeric wavelength filters are produced by a two-step molding process where the master mold is first formed on a negative tone photoresist and subsequently transferred to a PDMS mold; following this step, the PDMS silicon rubber mold was used as a stamp to transfer the pattern of the polymeric wavelength filters onto a UV cure epoxy.

Fabrication of polymer waveguides by a replication method.

We have developed a soft-lithography method to replicate polymer waveguides. In this method, the waveguides are produced by a two-step molding process where a master mold is first formed on a negative-tone photoresist and subsequently transferred to a polydimethylsiloxane (PDMS) mold; a PDMS silicone rubber mold is then used as a stamp to transfer the final waveguide pattern onto an UV cure epoxy. Initial results show good pattern transferring in physical shape.

Transducing mechanical force by use of a diffraction grating sensor.

A novel means of transducing mechanical force by using a polymeric-based diffractive grating sensor is presented. The diffraction gratings are successfully fabricated upon poly(dimethyl siloxane) polymer substrates by holographic interference and micromolding. A micromaterial tensile test incorporated into the surface diffraction grating experiment showed that a relationship between the load and the observed diffraction-pattern shift could be obtained.

Fabrication of a high-resolution periodical structure using a replication process.

We describe a procedure for rapidly and conveniently prototyping a periodic structure at submicrometer order using holographic interferometry and micro-molding processes. In this experiment, the master of the periodic structure was created on an i-line submicrometer positive photoresist film by a holographic interference using a He-Cd (325nm) laser. A subsequent mold using polydimethylsiloxane (PDMS) polymer was cast against this master and used as a stamp to transfer the grating pattern onto a UV cure epoxy.