BB rat Gimap gene expression in sorted lymphoid T and B cells

Aims: The Gimap gene family has been shown to be integral to T cell survival and development. A frameshift mutation in Gimap5, one of seven members of the Gimap family, results in lymphopenia and is a prerequisite for spontaneous type 1 diabetes (T1D) in the BioBreeding (BB) rat. While not contributing to lymphopenia, the Gimap family members proximal to Gimap5, encompassed within the Iddm39 quantitative trait locus (QTL), have been implicated in T1D. We hypothesized that expression of the Gimap family members within the Iddm39 QTL, during thymocyte development as well as in peripheral T and B cells contribute to T1D. Main methods: Cell sorted subpopulations were analyzed by quantitative real time (qRT) PCR. Key findings: Gimap4 expression was reduced in DR.(lyp/lyp) rat double negative, double positive and CD8 single positive (SP) thymocytes while expression of Gimap8, Gimap6, and Gimap7 was reduced only in CD8 SP thymocytes. Interestingly, expression of the entire Gimap gene family was reduced in DR.(lyp/lyp) rat peripheral T cells compared to non-lymphopenic, non-diabetic DR.(+/+) rats. With the exception of Gimap6. the Gimap family genes were not expressed in B cells from spleen and mesenteric lymph node (MLN). Expression of Gimap9 was only detected in hematopoietic cells of non B cell lineage such as macrophage, dendritic or NK cells. Significance: These results suggest that lack of the Gimap5 protein in the DR.(lyp/lyp) congenic rat was associated with impaired expression of the entire family of Gimap genes and may regulate T cell homeostasis in the peripheral lymphoid organs. (C) 2011 Elsevier Inc. All rights reserved

Cell-specific transcriptome changes in the hypothalamic arcuate nucleus in a mouse deoxycorticosterone acetate-salt model of hypertension

A common preclinical model of hypertension characterized by low circulating renin is the “deoxycorticosterone acetate (DOCA)-salt” model, which influences blood pressure and metabolism through mechanisms involving the angiotensin II type 1 receptor (AT1R) in the brain. More specifically, AT1R within Agouti-related peptide (AgRP) neurons of the arcuate nucleus of the hypothalamus (ARC) has been implicated in selected effects of DOCA-salt. In addition, microglia have been implicated in the cerebrovascular effects of DOCA-salt and angiotensin II. To characterize DOCA-salt effects upon the transcriptomes of individual cell types within the ARC, we used single-nucleus RNA sequencing (snRNAseq) to examine this region from male C57BL/6J mice that underwent sham or DOCA-salt treatment. Thirty-two unique primary cell type clusters were identified. Sub-clustering of neuropeptide-related clusters resulted in identification of three distinct AgRP subclusters. DOCA-salt treatment caused subtype-specific changes in gene expression patterns associated with AT1R and G protein signaling, neurotransmitter uptake, synapse functions, and hormone secretion. In addition, two primary cell type clusters were identified as resting versus activated microglia, and multiple distinct subtypes of activated microglia were suggested by sub-cluster analysis. While DOCA-salt had no overall effect on total microglial density within the ARC, DOCA-salt appeared to cause a redistribution of the relative abundance of activated microglia subtypes. These data provide novel insights into cell-specific molecular changes occurring within the ARC during DOCA-salt treatment, and prompt increased investigation of the physiological and pathophysiological significance of distinct subtypes of neuronal and glial cell types

Sequence Variation and Expression of the Gimap Gene Family in the BB Rat

Positional cloning of lymphopenia (lyp) in the BB rat revealed a frameshift mutation in Gimap5, a member of at least seven related GTPase Immune Associated Protein genes located on rat chromosome 4q24. Our aim was to clone and sequence the cDNA of the BB diabetes prone (DP) and diabetes resistant (DR) alleles of all seven Gimap genes in the congenic DR.lyp rat line with 2 Mb of BB DP DNA introgressed onto the DR genetic background. All (100%) DR.lyp/lyp rats are lymphopenic and develop type 1 diabetes (T1D) by 84 days of age while DR.+/+ rats remain T1D and lyp resistant. Among the seven Gimap genes, the Gimap5 frameshift mutation, a mutant allele that produces no protein, had the greatest impact on lymphopenia in the DR.lyp/lyp rat. Gimap4 and Gimap1 each had one amino acid substitution of unlikely significance for lymphopenia. Quantitative RT-PCR analysis showed a reduction in expression of all seven Gimap genes in DR.lyp/lyp spleen and mesenteric lymph nodes when compared to DR.+/+. Only four; Gimap1, Gimap4, Gimap5, and Gimap9 were reduced in thymus. Our data substantiates the Gimap5 frameshift mutation as the primary defect with only limited contributions to lymphopenia from the remaining Gimap genes

Genetic dissection reveals diabetes loci proximal to the gimap5 lymphopenia gene

rats are protected from type 1 diabetes (T1D) by 34 Mb of F344 DNA introgressed proximal to the gimap5 lymphopenia gene. To dissect the genetic factor(s) that confer protection from T1D in the DRF. f/f rat line, DRF. f/f rats were crossed to inbred BBDR or DR. lyp/lyp rats to generate congenic sublines that were genotyped and monitored for T1D, and positional candidate genes were sequenced. All (100%) DR. f/f congenic sublines further refined the RNO4 region 1 interval to ϳ670 kb and region 2 to the 340 kb proximal to gimap5. All congenic DRF. f/f sublines were prone to low-grade pancreatic mononuclear cell infiltration around ducts and vessels, but Ͻ20% of islets in nondiabetic rats showed islet infiltration. Coding sequence analysis revealed TCR V␤ 8E, 12, and 13 as candidate genes in region 1 and znf467 and atp6v0e2 as candidate genes in region 2. Our results show that spontaneous T1D is controlled by at least two genetic loci 7 Mb apart on rat chromosome 4. type 1 diabetes; BB rat; T cell receptor; autoimmune CHARACTERISTICS OF TYPE 1 DIABETES (T1D) in both human and the BioBreeding spontaneously diabetes-prone (BBDP) rat include polyuria, hyperglycemia, ketoacidosis, insulitis, and insulin dependency for life. As in human T1D, islets are infiltrated by mononuclear cells at the time of onset with rapid hyperglycemia due to a complete loss of islet ␤-cells (32). The genetic etiology of human T1D remains complex and although the major histocompatibility complex (MHC) (HLA DQ) on chromosome 6 accounts for ϳ40% of T1D risk, the number of non-HLA genetic factors is increasing steadily (2, 7). The BB rat offers a powerful model to dissect both genetic contributions and mechanisms by which immunemediated beta cell killing induces T1D (3, 4, 15, 17-21, 27, 28, 46). As in humans, the major genetic determinant of susceptibility in the BB rat is the MHC (Iddm1) on rat chromosome (RNO) 20. The class II MHC locus RT1B/D. u/u ), an ortholog of human HLA DQ (9), is necessary but not sufficient for T1D in the BBDP rat and other RT1. u/u -related rat strains with spontaneous (24, 47) or induced T1D (8, 43). In BBDP, a null mutation in the gimap5 gene (lyp; Iddm2) on RNO4 (14, 27) causes lymphopenia and is tightly linked to spontaneous T1D development. The DR. lyp/lyp rat with 2 Mb of BBDP DNA encompassing gimap5 introgressed into the genome of related BBDR rats (BioBreeding resistant to spontaneous T1D) are also 100% lymphopenic and 100% spontaneously diabetic (11). With complete T1D penetrance and tight regulation of onset, the congenic DR. lyp/lyp rat line offers distinct advantages in identification of genes responsible for disease progression. It is possible to induce T1D in BBDR rats (32) and related RT1 u/u rats (8) by administration of polyinosinic: polycytidylic acid (poly I:C, an activator of innate immunity), the T reg depleting cytotoxic DS4.23 anti-ART2.1 (formerly RT6) monoclonal antibody or by viral infection (34). This indicates that the BBDR has an underlying genetic susceptibility to T1D. In crosses between WF and either BBDP or BBDR rats, a quantitative trait locus (QTL) important for induced T1D (Iddm14, previously designated Iddm4) was mapped to RNO4 (6, Interestingly, F344 DNA introgressed between D4Rat253 and D4Rhw6 into the congenic DR. lyp/lyp genetic background resulted in a lymphopenic but nondiabetic rat (designated DRF. f/f ) (11). Protection from T1D in the DRF. f/f congenic rat line led us to conclude that spontaneous T1D in the BB rat is controlled, in part, by a diabetogenic factor(s) independent of the gimap5 mutation (76.84 Mb) on RNO4. This congenic interval is encompassed within Iddm14, raising the possibility that the Iddm14 locus could be required for both spontaneous and induced T1D in the BB rat. The aim of this study was to cross the DRF. f/f rat to BBDR and DR. lyp/lyp rats and produce recombinant sublines that could be assessed for both lymphopenia and diabetes and to estimate the number of independent genes on RNO4 that control spontaneous T1D

The genome sequence of the Norway rat, Rattus norvegicus Berkenhout 1769.

We present a genome assembly from an individual male Rattus norvegicus (the Norway rat; Chordata; Mammalia; Rodentia; Muridae). The genome sequence is 2.44 gigabases in span. The majority of the assembly is scaffolded into 20 chromosomal pseudomolecules, with both X and Y sex chromosomes assembled. This genome assembly, mRatBN7.2, represents the new reference genome for R. norvegicus and has been adopted by the Genome Reference Consortium

An international effort towards developing standards for best practices in analysis, interpretation and reporting of clinical genome sequencing results in the CLARITY Challenge

BACKGROUND: There is tremendous potential for genome sequencing to improve clinical diagnosis and care once it becomes routinely accessible, but this will require formalizing research methods into clinical best practices in the areas of sequence data generation, analysis, interpretation and reporting. The CLARITY Challenge was designed to spur convergence in methods for diagnosing genetic disease starting from clinical case history and genome sequencing data. DNA samples were obtained from three families with heritable genetic disorders and genomic sequence data was donated by sequencing platform vendors. The challenge was to analyze and interpret these data with the goals of identifying disease causing variants and reporting the findings in a clinically useful format. Participating contestant groups were solicited broadly, and an independent panel of judges evaluated their performance. RESULTS: A total of 30 international groups were engaged. The entries reveal a general convergence of practices on most elements of the analysis and interpretation process. However, even given this commonality of approach, only two groups identified the consensus candidate variants in all disease cases, demonstrating a need for consistent fine-tuning of the generally accepted methods. There was greater diversity of the final clinical report content and in the patient consenting process, demonstrating that these areas require additional exploration and standardization. CONCLUSIONS: The CLARITY Challenge provides a comprehensive assessment of current practices for using genome sequencing to diagnose and report genetic diseases. There is remarkable convergence in bioinformatic techniques, but medical interpretation and reporting are areas that require further development by many groups

The Gene Ontology knowledgebase in 2023

The Gene Ontology (GO) knowledgebase (http://geneontology.org) is a comprehensive resource concerning the functions of genes and gene products (proteins and noncoding RNAs). GO annotations cover genes from organisms across the tree of life as well as viruses, though most gene function knowledge currently derives from experiments carried out in a relatively small number of model organisms. Here, we provide an updated overview of the GO knowledgebase, as well as the efforts of the broad, international consortium of scientists that develops, maintains, and updates the GO knowledgebase. The GO knowledgebase consists of three components: (1) the GO-a computational knowledge structure describing the functional characteristics of genes; (2) GO annotations-evidence-supported statements asserting that a specific gene product has a particular functional characteristic; and (3) GO Causal Activity Models (GO-CAMs)-mechanistic models of molecular "pathways" (GO biological processes) created by linking multiple GO annotations using defined relations. Each of these components is continually expanded, revised, and updated in response to newly published discoveries and receives extensive QA checks, reviews, and user feedback. For each of these components, we provide a description of the current contents, recent developments to keep the knowledgebase up to date with new discoveries, and guidance on how users can best make use of the data that we provide. We conclude with future directions for the project