End-to-End Semi-automated Mid-scale Protein Screening Platform for Drug Discovery Research.

The drug discovery landscape is ever-evolving and constantly demands revolutionary technology advancements in protein expression and production laboratories. We have built a higher-throughput mid-scale semi-automated protein expression and screening platform to accelerate drug discovery research. The workflow described here enables comprehensive expression and purification screening assessment of challenging or difficult-to-express recombinant proteins in a fast and efficient manner by delivering small but sufficient amounts of high-quality proteins.

Restricted epitope specificity determined by variable region germline segment pairing in rodent antibody repertoires.

Antibodies from B-cell clonal lineages share sequence and structural properties as well as epitope specificity. Clonally unrelated antibodies can similarly share sequence and specificity properties and are said to be convergent. Convergent antibody responses against several antigens have been described in humans and mice and include different classes of shared sequence features.

Massively parallel single-cell B-cell receptor sequencing enables rapid discovery of diverse antigen-reactive antibodies.

Obtaining full-length antibody heavy- and light-chain variable regions from individual B cells at scale remains a challenging problem. Here we use high-throughput single-cell B-cell receptor sequencing (scBCR-seq) to obtain accurately paired full-length variable regions in a massively parallel fashion. We sequenced more than 250,000 B cells from rat, mouse and human repertoires to characterize their lineages and expansion.

Actionable Activating Oncogenic ERBB2/HER2 Transmembrane and Juxtamembrane Domain Mutations.

Deregulated HER2 is a target of many approved cancer drugs. We analyzed 111,176 patient tumors and identified recurrent mutations in HER2 transmembrane domain (TMD) and juxtamembrane domain (JMD) that include G660D, R678Q, E693K, and Q709L. Using a saturation mutagenesis screen and testing of patient-derived mutations we found several activating TMD and JMD mutations.

Comprehensive genomic analysis identifies pathogenic variants in maturity-onset diabetes of the young (MODY) patients in South India.

Background: Maturity-onset diabetes of the young (MODY) is an early-onset, autosomal dominant form of non-insulin dependent diabetes. Genetic diagnosis of MODY can transform patient management. Earlier data on the genetic predisposition to MODY have come primarily from familial studies in populations of European origin.

Massively parallel nanowell-based single-cell gene expression profiling.

Background: Technological advances have enabled transcriptome characterization of cell types at the single-cell level providing new biological insights. New methods that enable simple yet high-throughput single-cell expression profiling are highly desirable.

Results: Here we report a novel nanowell-based single-cell RNA sequencing system, ICELL8, which enables processing of thousands of cells per sample.

Phosphoregulatory protein 14-3-3 facilitates SAC1 transport from the endoplasmic reticulum.

Most secretory cargo proteins in eukaryotes are synthesized in the endoplasmic reticulum and actively exported in membrane-bound vesicles that are formed by the cytosolic coat protein complex II (COPII). COPII proteins are assisted by a variety of cargo-specific adaptor proteins required for the concentration and export of secretory proteins from the endoplasmic reticulum (ER). Adaptor proteins are key regulators of cargo export, and defects in their function may result in disease phenotypes in mammals.

Spectrum of diverse genomic alterations define non-clear cell renal carcinoma subtypes.

To further understand the molecular distinctions between kidney cancer subtypes, we analyzed exome, transcriptome and copy number alteration data from 167 primary human tumors that included renal oncocytomas and non-clear cell renal cell carcinomas (nccRCCs), consisting of papillary (pRCC), chromophobe (chRCC) and translocation (tRCC) subtypes. We identified ten significantly mutated genes in pRCC, including MET, NF2, SLC5A3, PNKD and CPQ. MET mutations occurred in 15% (10/65) of pRCC samples and included previously unreported recurrent activating mutations.

Ubiquitin-dependent regulation of COPII coat size and function.

Packaging of proteins from the endoplasmic reticulum into COPII vesicles is essential for secretion. In cells, most COPII vesicles are approximately 60-80 nm in diameter, yet some must increase their size to accommodate 300-400 nm procollagen fibres or chylomicrons. Impaired COPII function results in collagen deposition defects, cranio-lenticulo-sutural dysplasia, or chylomicron retention disease, but mechanisms to enlarge COPII coats have remained elusive.

The [corrected] SEC23-SEC31 [corrected] interface plays critical role for export of procollagen from the endoplasmic reticulum.

COPII proteins are essential for exporting most cargo molecules from the endoplasmic reticulum. The membrane-facing surface of the COPII proteins (especially SEC23-SEC24) interacts directly or indirectly with the cargo molecules destined for exit. As we characterized the SEC23A mutations at the SEC31 binding site identified from patients with cranio-lenticulo-sutural dysplasia, we discovered that the SEC23-SEC31 interface can also influence cargo selection.

COPII and the regulation of protein sorting in mammals.

Secretory proteins are transported to the Golgi complex in vesicles that bud from the endoplasmic reticulum. The cytoplasmic coat protein complex II (COPII) is responsible for cargo sorting and vesicle morphogenesis. COPII was first described in Saccharomyces cerevisiae, but its basic function is conserved throughout all eukaryotes.