Pavelec, D. | bioRxiv Science

bioRxiv2025

Caloric restriction reprograms adipose tissues in rhesus monkeys

Caloric restriction (CR) is a dietary intervention that delays the onset of age-related diseases and enhances survival in diverse organisms, and although changes in adipose tissues have been implicated in the beneficial effects of CR the molecular details are unknown. Here we show shared and depot-specific adaptations to life-long CR in subcutaneous and visceral adipose depots taken from advanced age male rhesus monkeys. Differential gene expression and pathway analysis identified key differences between the depots in metabolic, immune, and inflammatory pathways. In response to CR, RNA processing and proteostasis-related pathways were enriched in both depots but changes in metabolic, growth, and inflammatory pathways were depot-specific. Commonalities and differences that distinguish adipose depots are shared among monkeys and humans and the response to CR is highly conserved. These data reveal depot-specificity in adipose tissue adaptation that likely reflects differences in function and contribution to age-related disease vulnerability. In BriefRhesus monkey adipose tissues are highly responsive to long-term caloric restriction, although the response is depot-specific. Shared features include proteostasis and RNA processing pathways, whereas metabolic pathways, alterations in processes influenced by growth signaling, and changes in exon usage were depot-specific. The impact of CR on subcutaneous adipose tissue is highly conserved between monkeys and humans. HighlightsO_LIAt the transcript level, SAT and VAT taken from the same animals show modest specialization, primarily in metabolic and immune pathways. C_LIO_LIThe transcriptome of SAT is more responsive to CR than that of VAT, with depot-specific differences in metabolic adaptation and cell type composition. C_LIO_LIRNA processing pathways are engaged by CR and transcript isoforms are enriched in both depots, but differential exon usage is limited to SAT. C_LIO_LIAdipose reprogramming is linked to CR induced differences in body composition and systemic indices of health. C_LIO_LIThe impact of CR on subcutaneous adipose tissue is highly similar between monkeys and humans. C_LI

genomics↗

bioRxiv2022

A protocol for locating and counting transgenic sequences from laboratory animals using a map-then-capture (MapCap) sequencing workflow: procedure and application of results.

Transgenic rodent models for human diseases have been widely used over the past 50 years and are a mainstay of many biomedical research programs. Oftentimes the sequence of the transgenic segment of DNA is carefully designed but incorporation of this DNA into the host genome is less well understood. Structural variation and insertional mutagenesis may occur at transgenic insertion sites. Here, we present a robust workflow including identification of the transgene locus via selective Illumina sequencing followed by Cas9-mediated target DNA enrichment of the locus, which successfully identified beginning and end sites of a large transgenic insertion into a murine model for human amylin-induced type II diabetes. Enriched sequences were mapped via Oxford Nanopore sequencing. Although the insertion was too long for a single mapped genetic sequence to encompass, the method provided multiple insights relevant to the animal model: a minimum number of forward- and reverse-facing transcript copies as well as characterization of an inversion point within the insertion site. The insertion start point containing both murine and human DNA was used to identify and separate animals hemizygous for the transgenic insertion from homozygous animals. This identification could be performed early in the rodent life cycle prior to maturation (i.e. breeding age), thus allowing for management of colony phenotypes and eliminating the need to "genotype by phenotype" later on (onset of amylin-induced type II diabetes does not occur until ~8-10 weeks of age for this model). We further confirmed our homozygous diabetic mice function the same as colonies established in other labs and present full antibody and fluorescent-staining protocols (available in SI). Lastly, we note that, due to our genotyping, a novel animal was identified within our colony: non-diabetic homozygous mice. Indeed, only 37% of homozygous mice bred in our colony became diabetic. AUTHOR SUMMARY (for broad audience)Transgenic rodent models are important to studying human diseases. When creating a new rodent model, one may insert new DNA into a well-characterized background genome. However, it is oftentimes not known where the new DNA was incorporated, how many times it was incorporated, or if any coding sequences or regulatory elements within native DNA were disrupted. Here, we have developed a method to characterize transgenic animals, and have applied it to a popular model for studying human amylin-induced type II diabetes.

genetics↗