Â鶹´«Ã½É«ÇéƬ

We offer a selection of recent papers on a variety of topics from the Journal of Biological Chemistry, the Journal of Lipid Research and Â鶹´«Ã½É«ÇéƬ & Cellular Proteomics.

Blood and aggression in crayfish

Transglutaminase is a cross-linking enzyme involved in diverse cellular functions including blood coagulation and communication between cells and the extracellular matrix. Different isoforms of the enzyme have different substrate specificities and are expressed differentially across tissues and cell types. Kingkamon Junkunlo and colleagues at Uppsala University examined the role of transglutaminase in hematopoiesis in crayfish. They found that inhibiting transglutaminase not only increased the number of circulating blood cells but also unexpectedly made crayfish move less and display less aggressive behavior. This link between hematopoiesis and the nervous system controlling behavior may provide insights into diseases involving defects in motor functions. was published in the Journal of Biological Chemistry.

Early markers of Alzheimer’s disease

Alzheimer’s disease is characterized by neurodegeneration, including synapse loss, and preclinical markers of Alzheimer’s and other neuronal disorders have been elusive. These markers someday could be used to diagnose patients before symptoms start.

Alberto Lleo, Olivia Belbin and colleagues at the Hospital de la Santa Creu i Sant Pau in Barcelona, Spain, used a mass spectrometry-based proteomic technique to determine which proteins in cerebrospinal fluid, or CSF, samples change prior to the onset of Alzheimer’s symptoms. were published in the journal Â鶹´«Ã½É«ÇéƬ & Cellular Proteomics.

The researchers first compared CSF proteins in Alzheimer’s patients to those in people without the disease. Next, they narrowed the set down to CSF proteins from synapses, identified by comparing the data set to synaptic proteins established in the literature. Nine were chosen for monitoring changes in expression levels in Alzheimer’s patients.

Using a proteomics technique called selected reaction monitoring, the researchers found that while some marker proteins were high throughout the onset of disease, several proteins were lower in CSF from presymptomatic patients than in healthy individuals but then were elevated where neurodegeneration occurred. Lleo and colleagues described this as a “biphasic profile” and concluded that those decreased proteins could be early biomarkers of synapse loss that could be used to identify individuals with Alzheimer’s disease before irreversible neurodegeneration becomes widespread.

The new old cause of gallstones

A person living in the U.S. has a 10 to 20 percent chance of developing stones in their gallbladder. Yet up to 80 percent of affected individuals will have no symptoms. The remaining 20 percent may experience biliary pain or other complications requiring costly interventions.

In in the Journal of Lipid Research, Mats Rudling and colleagues at the Karolinska Institute, Sweden, describe new insight into the cause of gallstones.

The stones develop when gallbladder bile is supersaturated with cholesterol, one of three major lipid components in bile, with bile acids and phospholipids being the other two. Traditionally, it is thought that excessive hepatic secretion of cholesterol is the main cause for such supersaturation. Rudling’s team used unpublished data from a previous study to show that a shortage of bile acids is probably the major and most common reason why gallbladder bile is supersaturated with cholesterol in gallstone patients. Furthermore, the researchers used 13 previously published studies from 11 countries to corroborate their findings. Future studies are needed to investigate the cause(s) of reduced bile acid levels.

Discovering a new pool of iron

Like many elements, iron is a double-edged sword. It is an essential cofactor but can pose a danger to cells by catalyzing the formation of reactive oxygen species. In in the Journal of Biological Chemistry, Joshua Wofford and colleagues at Texas A&M University describe how they revised previous estimates of iron usage in cells by using multiple methods to inventory total iron content and characterize the distribution of different iron species in Escherichia coli. They discovered a previously unknown low-molecular-mass Fe(II) pool in the cytosol and suggested that this pool might be prevented from reacting with oxygen by a shield of membrane-bound iron-rich respiratory complexes.

DNA repair pathway resolved

DNA repair is a critical function, and cells have multiple ways of carrying it out. One efficient method of DNA repair is transcription-coupled repair, which is carried out as DNA is being transcribed. Previous research provided conflicting evidence as to whether transcription-coupled repair worked on DNA sequences encoding ribosomal RNA; this was unclear because rRNA and protein-coding genes are transcribed by different RNA polymerases in eukaryotes. Yanyan Yang and colleagues at the University of North Carolina used excision repair sequencing with single-nucleotide resolution to investigate this question. They found that transcribed and non-transcribed sections of ribosomal DNA are repaired at the same rates, suggesting that ribosomal DNA is repaired by global repair mechanisms, not transcription-coupled repair. was published in the Journal of Biological Chemistry.

Glycoproteins and food poisoning

The gastrointestinal bacterium Campylobacter jejuni, a major cause of food poisoning, causes abdominal cramps and inflammation and is transmitted through contaminated water or poultry. C. jejuni uses a sophisticated system to attach oligosaccharide chains to the nitrogen in some amino acids. The system involves multiple steps carried out by the pgl cluster, which codes for proteins that act as a glycoprotein assembly line. Researchers at the University of Sydney generated C. jejuni bacteria without a key gene in this cluster and performed proteomic analysis to determine the importance of N-glycosylation in pathogenesis. was published in the journal Â鶹´«Ã½É«ÇéƬ & Cellular Proteomics.

Researchers found that without the pglB protein, C. jejuni no longer could recover from extreme cold or heat as normal bacteria could. In addition, C. jejuni switched which amino acids it used most, decreased levels of certain transport proteins and could no longer migrate toward some metabolic substrates.

Without glycosylation, certain proteins within the glycoproteome were downregulated, and an enzyme involved in utilization of nitrogen was less active. This was specific; a strain without the pgl cluster had restored activity of this enzyme when researchers added the pgl cluster back. Because protein glycosylation in C. jejuni was found to affect many pathways, it could be a way to stop this gastrointestinal bacterium.

How an upstream enhancer regulates a protein’s expression

GPIHBP1 is a protein that shuttles lipoprotein lipase, or LPL, from capillary endothelial cells to capillary lumen, where it can process triglyceride-rich lipoproteins. Lack of GPIHBP1 prevents LPL relocation to the capillary lumen, impairing plasma triglyceride hydrolysis and leading to severe hypertriglyceridemia (high blood levels of triglycerides — a risk factor for atherosclerosis).

In published in the Journal of Lipid Research, Christopher Allan and colleagues at the universities of California, Michigan and Arizona describe an upstream enhancer that regulates GPIHBP1 expression in a tissue-specific manner. Deletion of the relevant DNA segment using CRISPR/Cas9 genome editing reduced GPIHBP1 expression by more than 90 percent in the liver and by about 50 percent in the heart and brown adipose tissue, but that was wasn’t enough to block LPL relocalization to the capillary lumen and cause hypertriglyceridemia. Future studies may investigate why the enhancer deletion had such strikingly different effects in the liver versus the heart and focus on other unknown players in the regulation of GPIHBP1 expression. It is likely that other GPIHBP1 enhancers were present, which contributed to the tissue-specific changes in expression.

Finding ligands using phage

USP11 is a human deubiquitinase that regulates DNA double-strand break repair and cellular stress responses; it is dysregulated in pancreatic and ovarian cancers. Unlike other ubiquitin-specific proteases, USP11’s peptide-binding sites were unknown. Anastasios Spiliotopoulos and colleagues at the University of Nottingham combined phage display library screening with next-generation sequencing to discover USP11-interacting peptide motifs. They found a binding site that modulates USP11’s DNA repair function and developed a ligand that could be used to arrest the cell cycle. This strategy, in the Journal of Biological Chemistry, could be used to develop new biochemical tools.

Selectivity hinges on a hinge site

Despite being structurally similar, caspases are highly selective for specific substrates. In published in the Journal of Biological Chemistry, Derek McPherson and colleagues at the University of Massachusetts searched for secondary binding sites in caspases that could help explain this selectivity. They discovered an exosite in caspase-6 composed of a tri-arginine patch in the hinge between the core caspase domain and a disordered N-terminal domain. Since caspase-6 plays a unique role among apoptotic caspases in neurodegeneration, modulating this substrate-binding site could be therapeutically valuable for conditions such as Alzheimer’s and Huntington’s disease.