Genital endometriosis: controversial issues and alternative approaches to diagnosis and treatment

On the basis of the Department of Operative Gynecology of the Scientific Center for Obstetrics, Gynecology and Perinatology of the Russian Academy of Medical Sciences from 1992 to 2002. examination, treatment and monitoring of 1234 patients with endometriosis was carried out, which served as material for the study of morphofunctional, endocrine, immune, biochemical, genetic aspects of endometriosis. Our clinical and scientific experience allowed us to determine our own positions and a circle of discussion questions corresponding to the main directions of further research on this problem, which are: is endometriosis a disease or not ?; development mechanisms; classification; diagnostic criteria; features of histological characteristics; endometriosis and genetics; endometriosis and the immune system; endometriosis and adenomyosis; retrocervical endometriosis; endometriosis and pelvic pain; endometriosis and adhesions; endometriosis and infertility; traditional and non-traditional approaches to diagnosis and treatment.

Exploring the Diversity and Regulation of Apocarotenoid Metabolic Pathways in Plants

In plants, carotenoids are subjected to enzyme-catalyzed oxidative cleavage reactions as well as to non-enzymatic degradation processes, which produce various carbonyl products called apocarotenoids. These conversions control carotenoid content in different tissues and give rise to apocarotenoid hormones and signaling molecules, which play important roles in plant growth and development, response to environmental stimuli, and in interactions with surrounding organisms. In addition, carotenoid cleavage gives rise to apocarotenoid pigments and volatiles that contribute to the color and flavor of many flowers and several fruits. Some apocarotenoid pigments, such as crocins and bixin, are widely utilized as colorants and additives in food and cosmetic industry and also have health-promoting properties. Considering the importance of this class of metabolites, investigation of apocarotenoid diversity and regulation has increasingly attracted the attention of plant biologists. Here, we provide an update on the plant apocarotenoid biosynthetic pathway, especially highlighting the diversity of the enzyme carotenoid cleavage dioxygenase 4 (CCD4) from different plant species with respect to substrate specificity and regioselectivity, which contribute to the formation of diverse apocarotenoid volatiles and pigments. In addition, we summarize the regulation of apocarotenoid metabolic pathway at transcriptional, post-translational, and epigenetic levels. Finally, we describe inter- and intraspecies variation in apocarotenoid production observed in many important horticulture crops and depict recent progress in elucidating the genetic basis of the natural variation in the composition and amount of apocarotenoids. We propose that the illustration of biochemical, genetic, and evolutionary background of apocarotenoid diversity would not only accelerate the discovery of unknown biosynthetic and regulatory genes of bioactive apocarotenoids but also enable the identification of genetic variation of causal genes for marker-assisted improvement of aroma and color of fruits and vegetables and CRISPR-based next-generation metabolic engineering of high-value apocarotenoids.

Disassembling the Nature of Capsid: Biochemical, Genetic, and Imaging Approaches to Assess HIV-1 Capsid Functions

The human immunodeficiency virus type 1 (HIV-1) capsid and its disassembly, or capsid uncoating, has remained an active area of study over the past several decades. Our understanding of the HIV-1 capsid as solely a protective shell has since shifted with discoveries linking it to other complex replication events. The interplay of the HIV-1 capsid with reverse transcription, nuclear import, and integration has led to an expansion of knowledge of capsid functionality. Coincident with advances in microscopy, cell, and biochemistry assays, several models of capsid disassembly have been proposed, in which it occurs in either the cytoplasmic, nuclear envelope, or nuclear regions of the cell. Here, we discuss how the understanding of the HIV-1 capsid has evolved and the key methods that made these discoveries possible.

Nuclear export of the pre-60S ribosomal subunit through single nuclear pores observed in real time

Ribosomal biogenesis has been studied by biochemical, genetic and electron microscopic approaches, but live cell data on the in vivo kinetics are still missing. Here we analyse the export kinetics of the large ribosomal subunit (pre-60S particle) through single NPCs in human cells. We established a stable cell line co-expressing Halo-tagged eIF6 and GFP-fused NTF2 to simultaneously label pre-60S particles and NPCs, respectively. By combining single molecule tracking and super resolution confocal microscopy we visualize the dynamics of single pre-60S particles during export through single NPCs. For export events, maximum particle accumulation is found in the centre of the pore, while unsuccessful export terminates within the nuclear basket. The export has a single rate limiting step and a duration of ∼24 milliseconds. Only about 1/3 of attempted export events are successful. Our results show that the mass flux through a single NPC can reach up to ~125 MDa·s−1 in vivo.

Cryo EM Analysis Reveals Inherent Flexibility of Authentic Murine Papillomavirus Capsids

Human papillomavirus (HPV) is a significant health burden and leading cause of virus-induced cancers. However, studies have been hampered due to restricted tropism that makes production and purification of high titer virus problematic. This issue has been overcome by developing alternative HPV production methods such as virus-like particles (VLPs), which are devoid of a native viral genome. Structural studies have been limited in resolution due to the heterogeneity, fragility, and stability of the VLP capsids. The mouse papillomavirus (MmuPV1) presented here has provided the opportunity to study a native papillomavirus in the context of a common laboratory animal. Using cryo EM to solve the structure of MmuPV1, we achieved 3.3 Å resolution with a local symmetry refinement method that defined smaller, symmetry related subparticles. The resulting high-resolution structure allowed us to build the MmuPV1 asymmetric unit for the first time and identify putative L2 density. We also used our program ISECC to quantify capsid flexibility, which revealed that capsomers move as rigid bodies connected by flexible linkers. The MmuPV1 flexibility was comparable to that of a HPV VLP previously characterized. The resulting MmuPV1 structure is a promising step forward in the study of papillomavirus and will provide a framework for continuing biochemical, genetic, and biophysical research for papillomaviruses.

Blocking a dead-end assembly pathway creates a point of regulation for the DEAD-box ATPase Has1 and prevents platform misassembly

Assembly of ribosomal subunits occurs via parallel pathways, which accelerate the process and render it more robust. Nonetheless, in vitro analyses have also demonstrated that some assembly pathways are dead-ends, presumably due to rRNA misfolding. If and how these non-productive pathways are avoided during assembly in vivo remains unknown. Here we use a combination of biochemical, genetic, proteomic and structural analyses to demonstrate a role for assembly factors in biasing the folding landscape away from non-productive intermediates. By binding Rrp36, Rrp5 is prevented from forming a premature interaction with the platform, which leads to a dead-end intermediate, and a misassembled platform that is functionally defective. The DEAD-box ATPase Has1 separates Rrp5 and Rrp36, allowing Rrp5 to reposition to the platform, thereby promoting ribosome assembly and enabling rRNA processing. Thus, Rrp36 establishes an ATP-dependent regulatory point that ensures correct platform assembly by opening a new folding channel that avoids funnels to misfolding.

Bacterial translation machinery for deliberate mistranslation of the genetic code

Inaccurate expression of the genetic code, also known as mistranslation, is an emerging paradigm in microbial studies. Growing evidence suggests that many microbial pathogens can deliberately mistranslate their genetic code to help invade a host or evade host immune responses. However, discovering different capacities for deliberate mistranslation remains a challenge because each group of pathogens typically employs a unique mistranslation mechanism. In this study, we address this problem by studying duplicated genes of aminoacyl-transfer RNA (tRNA) synthetases. Using bacterial prolyl-tRNA synthetase (ProRS) genes as an example, we identify an anomalous ProRS isoform, ProRSx, and a corresponding tRNA, tRNAProA, that are predominately found in plant pathogens from Streptomyces species. We then show that tRNAProA has an unusual hybrid structure that allows this tRNA to mistranslate alanine codons as proline. Finally, we provide biochemical, genetic, and mass spectrometric evidence that cells which express ProRSx and tRNAProA can translate GCU alanine codons as both alanine and proline. This dual use of alanine codons creates a hidden proteome diversity due to stochastic Ala→Pro mutations in protein sequences. Thus, we show that important plant pathogens are equipped with a tool to alter the identity of their sense codons. This finding reveals the initial example of a natural tRNA synthetase/tRNA pair for dedicated mistranslation of sense codons.

Cytogenetic and Biochemical Genetic Techniques for Personalized Drug Therapy in Europe

For many authorized drugs, accumulating scientific evidence supports testing for predictive biomarkers to apply personalized therapy and support preventive measures regarding adverse drug reactions and treatment failure. Here, we review cytogenetic and biochemical genetic testing methods that are available to guide therapy with drugs centrally approved in the European Union (EU). We identified several methods and combinations of techniques registered in the Genetic Testing Registry (GTR), which can be used to guide therapy with drugs for which pharmacogenomic-related information is provided in the European public assessment reports. Although this registry provides information on genetic tests offered worldwide, we identified limitations regarding standard techniques applied in clinical practice and the information on test validity rarely provided in the according sections.