Ancient Faith Radio and Bill Marianes were honored to be present at the recent meeting of the Assembly of Bishops. This week, here Bill's interviews with Archbishop Elpidophoros of the Greek Orthodox Archdiocese, Metropolitan Joseph of the Antiochian Archdiocese, Metropolitan Tikhon of the Orthodox Church in America, Serbian Orthodox Bishop Irinej of Eastern America, Metropolitan Methodios of the Greek Orthodox Metropolis of Boston, Archbishop Daniel of the Ukranian Orthodox Church of the USA, and Metropolitan Savas of the Greek Orthodox Metropolis of Pittsburgh

Complex I (proton-pumping NADH:ubiquinone oxidoreductase) is the first component of the mitochondrial respiratory chain. In recent years, high-resolution cryo-EM studies of complex I from various species have greatly enhanced the understanding of the structure and function of this important membrane-protein complex. Less well studied is the structural basis of complex I biogenesis. The assembly of this complex of more than 40 subunits, encoded by nuclear or mitochondrial DNA, is an intricate process that requires at least 20 different assembly factors in humans. These are proteins that are transiently associated with building blocks of the complex and are involved in the assembly process, but are not part of mature complex I. Although the assembly pathways have been studied extensively, there is limited information on the structure and molecular function of the assembly factors. Here, the insights that have been gained into the assembly process using cryo-EM are reviewed.

The 12th Annual Assembly Show wrapped up at the Donald E. Stephens Convention Center in Rosemont, IL, with the announcement of the New Products Award winners.

Leamington’s finest opens its doors once more

There’s a definite eighties feel in Leamington.

Icons, influences and young whippersnappers.

After a few quiet weeks Leamington’s finest is springing back into life.

Archan Chakraborty, Wei-Cheng Lin, Yu-Tsun Lin, Kuang-Jing Huang, Pei-Yu Wang, Yi-Feng Chang, Hsiang-Iu Wang, Kung-Ting Ma, Chun-Yen Wang, Xuan-Rong Huang, Yen-Hsien Lee, Bi-Chang Chen, Ya-Ju Hsieh, Kun-Yi Chien, Tzu-Yang Lin, Ji-Long Liu, Li-Ying Sung, Jau-Song Yu, Yu-Sun Chang, and Li-Mei Pai

Under metabolic stress, cellular components can assemble into distinct membraneless organelles for adaptation. One such example is cytidine 5'-triphosphate synthase (CTPS), which forms filamentous structures under glutamine deprivation. We have previously demonstrated that histidine (His)-mediated methylation regulates the formation of CTPS filaments to suppress enzymatic activity and preserve the CTPS protein under Gln deprivation, which promotes cancer cell growth after stress alleviation. However, it remains unclear where and how these enigmatic structures are assembled. Using CTPS-APEX2-mediated in vivo proximity labeling, we found that SNAP29 regulates the spatiotemporal filament assembly of CTPS along the cytokeratin network in a keratin 8 (KRT8)-dependent manner. Knockdown of synaptosome-associated protein 29 (SNAP29) interfered with assembly and relaxed the filament-induced suppression of CTPS enzymatic activity. Furthermore, APEX2 proximity labeling of keratin 18 (KRT18) revealed a spatiotemporal association of SNAP29 with cytokeratin in response to stress. Super-resolution imaging suggests that during CTPS filament formation, SNAP29 interacts with CTPS along the cytokeratin network. This study links the cytokeratin network to the regulation of metabolism by compartmentalization of metabolic enzymes during nutrient deprivation.

Laurence Haren, Dorian Farache, Laurent Emorine, and Andreas Merdes

-tubulin is a major protein involved in the nucleation of microtubules in all eukaryotes. It forms two different complexes with proteins of the GCP family (gamma-tubulin complex proteins): -tubulin small complexes (TuSCs), containing -tubulin and GCPs 2 and 3, and -tubulin ring complexes (TuRCs), containing multiple TuSCs, in addition to GCPs 4, 5, and 6. Whereas the structure and assembly properties of TuSCs have been intensively studied, little is known about the assembly of TuRCs, and about the specific roles of GCPs 4, 5, and 6. Here, we demonstrate that two copies of GCP4 and one copy each of GCP5 and GCP6 form a salt-resistant sub-complex within the TuRC that assembles independently of the presence of TuSCs. Incubation of this sub-complex with cytoplasmic extracts containing TuSCs leads to the reconstitution of TuRCs that are competent to nucleate microtubules. In addition, we investigate sequence extensions and insertions that are specifically found at the amino-terminus of GCP6, and between the GCP6 grip1 and grip2 motifs, and we demonstrate that these are involved in the assembly or stabilization of the TuRC.

A flush adaptor for use with a valve fitment assembly for dispensing liquids from a container; wherein the flush adaptor comprises an outer ring-collar; a flange with an edge molded to the bottom of the outer ring-collar; an interior ring-collar adjacent to the outer ring-collar; a ridge molded in the interior ring-collar; a seat molded onto the interior ring-collar and a pin molded into the interior ring-collar which keeps the valve in an open position; and a hollow tube molded into the adaptor to allow the flow of liquid through the adaptor and into the fitment assembly; whereby the flush adaptor allows for cleaning of the assembly and any tubes connected thereto.

In conventional aircraft cargo compartments panels or similar flat floor elements are fastened to floor beams or similar supporting elements that are installed in the body of the aircraft. Subsequently functional units such as roller elements, latches or PDUs are mounted and connected to one another by way of appropriate control conductors. It is proposed to fasten the floor elements permanently to the supporting beams so as to form prefabricated floor modules and to install these floor modules in the aircraft.

apoB exists as apoB100 and apoB48, which are mainly found in hepatic VLDLs and intestinal chylomicrons, respectively. Elevated plasma levels of apoB-containing lipoproteins (Blps) contribute to coronary artery disease, diabetes, and other cardiometabolic conditions. Studying the mechanisms that drive the assembly, intracellular trafficking, secretion, and function of Blps remains challenging. Our understanding of the intracellular and intraorganism trafficking of Blps can be greatly enhanced, however, with the availability of fusion proteins that can help visualize Blp transport within cells and between tissues. We designed three plasmids expressing human apoB fluorescent fusion proteins: apoB48-GFP, apoB100-GFP, and apoB48-mCherry. In Cos-7 cells, transiently expressed fluorescent apoB proteins colocalized with calnexin and were only secreted if cells were cotransfected with microsomal triglyceride transfer protein. The secreted apoB-fusion proteins retained the fluorescent protein and were secreted as lipoproteins with flotation densities similar to plasma HDL and LDL. In a rat hepatoma McA-RH7777 cell line, the human apoB100 fusion protein was secreted as VLDL- and LDL-sized particles, and the apoB48 fusion proteins were secreted as LDL- and HDL-sized particles. To monitor lipoprotein trafficking in vivo, the apoB48-mCherry construct was transiently expressed in zebrafish larvae and was detected throughout the liver. These experiments show that the addition of fluorescent proteins to the C terminus of apoB does not disrupt their assembly, localization, secretion, or endocytosis. The availability of fluorescently labeled apoB proteins will facilitate the exploration of the assembly, degradation, and transport of Blps and help to identify novel compounds that interfere with these processes via high-throughput screening.

Last week, LM Wind Power said it had successfully manufactured the world’s first wind turbine blade to surpass 100 meters in length.