Multiple expressed MHC class II loci in salmonids; details of one non-classical region in Atlantic salmon (Salmo salar)

<p>Abstract</p> <p>Background</p> <p>In teleosts, the Major Histocompatibility Complex (MHC) class I and class II molecules reside on different linkage groups as opposed to tetrapods and shark, where the class I and class II genes reside in one genomic region. Several teleost MHC class I regions have been sequenced and show varying number of class I genes. Salmonids have one major expressed MHC class I locus (UBA) in addition to varying numbers of non-classical genes. Two other more distant lineages are also identifyed denoted L and ZE. For class II, only one major expressed class II alpha (DAA) and beta (DAB) gene has been identified in salmonids so far.</p> <p>Results</p> <p>We sequenced a genomic region of 211 kb encompassing divergent MHC class II alpha (<it>Sasa-DBA</it>) and beta (<it>Sasa-DBB</it>) genes in addition to NRGN, TIPRL, TBCEL and TECTA. The region was not linked to the classical class II genes and had some synteny to genomic regions from other teleosts. Two additional divergent and expressed class II sequences denoted DCA and DDA were also identified in both salmon and trout. Expression patterns and lack of polymorphism make these genes non-classical class II analogues. <it>Sasa-DBB</it>, <it>Sasa-DCA </it>and <it>Sasa-DDA </it>had highest expression levels in liver, hindgut and spleen respectively, suggestive of distinctive functions in these tissues. Phylogenetic studies revealed more yet undescribed divergent expressed MHC class II molecules also in other teleosts.</p> <p>Conclusion</p> <p>We have characterised one genomic region containing expressed non-classical MHC class II genes in addition to four other genes not involved in immune function. Salmonids contain at least two expressed MHC class II beta genes and four expressed MHC class II alpha genes with properties suggestive of new functions for MHC class II in vertebrates. Collectively, our data suggest that the class II is worthy of more elaborate studies also in other teleost species.</p

Genomic Organization of Duplicated Major Histocompatibility Complex Class I Regions in Atlantic Salmon (Salmo Salar)

Background: We have previously identified associations between major histocompatibility complex(MHC) class I and resistance towards bacterial and viral pathogens in Atlantic salmon. To evaluate if onlyMHC or also closely linked genes contributed to the observed resistance we ventured into sequencing ofthe duplicated MHC class I regions of Atlantic salmon.Results: Nine BACs covering more than 500 kb of the two duplicated MHC class I regions of Atlanticsalmon were sequenced and the gene organizations characterized. Both regions contained the proteasomecomponents PSMB8, PSMB9, PSMB9-like and PSMB10 in addition to the transporter for antigen processingTAP2, as well as genes for KIFC1, ZBTB22, DAXX, TAPBP, BRD2, COL11A2, RXRB and SLC39A7. TheIA region contained the recently reported MHC class I Sasa-ULA locus residing approximately 50 kbupstream of the major Sasa-UBA locus. The duplicated class IB region contained an MHC class I locusresembling the rainbow trout UCA locus, but although transcribed it was a pseudogene. No other MHCclass I-like genes were detected in the two duplicated regions. Two allelic BACs spanning the UBA locushad 99.2% identity over 125 kb, while the IA region showed 82.5% identity over 136 kb to the IB region.The Atlantic salmon IB region had an insert of 220 kb in comparison to the IA region containing threechitin synthase genes.Conclusion: We have characterized the gene organization of more than 500 kb of the two duplicatedMHC class I regions in Atlantic salmon. Although Atlantic salmon and rainbow trout are closely related,the gene organization of their IB region has undergone extensive gene rearrangements. The Atlanticsalmon has only one class I UCA pseudogene in the IB region while trout contains the four MHC UCA, UDA,UEA and UFA class I loci. The large differences in gene content and most likely function of the salmon andtrout class IB region clearly argues that sequencing of salmon will not necessarily provide informationrelevant for trout and vice versa

Comprehensive analysis of MHC class I genes from the U-, S-, and Z-lineages in Atlantic salmon

<p>Abstract</p> <p>Background</p> <p>We have previously sequenced more than 500 kb of the duplicated MHC class I regions in Atlantic salmon. In the IA region we identified the loci for the MHC class I gene <it>Sasa-UBA </it>in addition to a soluble MHC class I molecule, <it>Sasa-ULA</it>. A pseudolocus for <it>Sasa-UCA </it>was identified in the nonclassical IB region. Both regions contained genes for antigen presentation, as wells as orthologues to other genes residing in the human MHC region.</p> <p>Results</p> <p>The genomic localisation of two MHC class I lineages (Z and S) has been resolved. 7 BACs were sequenced using a combination of standard Sanger and 454 sequencing. The new sequence data extended the IA region with 150 kb identifying the location of one Z-lineage locus, <it>ZAA</it>. The IB region was extended with 350 kb including three new Z-lineage loci, <it>ZBA</it>, <it>ZCA </it>and <it>ZDA </it>in addition to a <it>UGA </it>locus. An allelic version of the IB region contained a functional <it>UDA </it>locus in addition to the <it>UCA </it>pseudolocus. Additionally a BAC harbouring two MHC class I genes (UHA) was placed on linkage group 14, while a BAC containing the S-lineage locus <it>SAA </it>(previously known as <it>UAA</it>) was placed on LG10. Gene expression studies showed limited expression range for all class I genes with exception of <it>UBA </it>being dominantly expressed in gut, spleen and gills, and <it>ZAA </it>with high expression in blood.</p> <p>Conclusion</p> <p>Here we describe the genomic organization of MHC class I loci from the U-, Z-, and S-lineages in Atlantic salmon. Nine of the described class I genes are located in the extension of the duplicated IA and IB regions, while three class I genes are found on two separate linkage groups. The gene organization of the two regions indicates that the IB region is evolving at a different pace than the IA region. Expression profiling, polymorphic content, peptide binding properties and phylogenetic relationship show that Atlantic salmon has only one MHC class Ia gene (<it>UBA</it>), in addition to a multitude of nonclassical MHC class I genes from the U-, S- and Z-lineages.</p

AI is a viable alternative to high throughput screening: a 318-target study

: High throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery

Guidelines for the use and interpretation of assays for monitoring autophagy

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field