The Peters Lab

Genetics of Blood Cell Development and Disease

Our current projects focus on three major themes. First is the identification of genetic modifiers of embryonic globin expression, critical to sickle cell disease therapy. Second is analysis of aberrant Rasa3-Ras function leading to bone marrow failure in the mutant mouse model, scat (severe combined anemia and thrombocytopenia). Third is the anemia of aging, which affects more than 3 million people 65 years of age or older in the United States (2009 data), is highly influenced by heredity, and is associated with multiple adverse outcomes.

Genetic modifiers of embryonic globin expression

During human development fetal hemoglobin (HbF), composed of two α- and two γ-globin chains (α₂γ₂), is down-regulated as erythropoiesis switches from the fetal liver to the bone marrow. In the bone marrow, β-globin expression increases and replaces γ, producing adult HbA (α₂β₂). In SCD, a Glu -> Val substitution in the 6^th residue of the adult β-globin chain produces sickle hemoglobin, HbS (α₂β^s₂). As HbF declines during late fetal development and the first few months of life, symptoms of SCD emerge as the ameliorative effects of HbF are lost. In mice, “maturational” globin switching occurs wherein βh1 embryonic globin expression switches to εγ as primitive red cells mature in the circulation. Postnatal elevation of HbF expression via co-inheritance of positive genetic modifiers or hydroxyurea (HU) treatment alleviate SCD severity. However, HU often has adverse side effects, including myelosuppression. Therefore, identification of novel drug targets allowing HbF up-regulation in patients is urgently needed. Although differences exist, many mechanisms of hemoglobin regulation are conserved between mice and humans.

To identify novel genetic modifiers of globin switching, we are mapping quantitative trait locus (QTL) for βh1 expression in F2 intercrosses using the Klf1^Nan mouse mutant. Klf^Nan carries a missense mutation in the gene encoding the KLF1 transcription factor that causes anemia throughout life and that is accompanied by dramatically increased expression of embryonic βh1 globin in adult erythroid cells. We are also mapping QTL for βh1 expression in the high resolution Diversity Outbred (DO) mouse population. These data will be integrated with RNAseq and chromatin immunoprecipitation (ChIP) assays to identify candidate genes. Our ultimate aim is to identify new therapeutic targets for drug discovery.

Analysis of aberrant Rasa3-Ras function

We recently described the inbred, autosomal recessive mouse mutant scat (severe combined anemia and thrombocytopenia). We demonstrated that scat carries a mutation in a gene not previously recognized as critical in vertebrate hematopoiesis, Rasa3. Scat is an intriguing, unique model characterized by bone marrow failure. Significantly, the disease is episodic. Homozygotes are severely affected in utero and at birth. A remarkable, spontaneous remission ensues in those surviving this first “crisis” period wherein all aspects of the disease phenotype disappear. A second crisis invariably follows, however, and 90% of homozygous mice die by 4-5 weeks of age due to catastrophic hematopoietic failure in both the bone marrow and spleen. Notably, bone marrow failure in humans is known to remit spontaneously as well. The remarkable reversion of the abnormal phenotype to normal in scat offers an entry point into gene networks relevant to bone marrow failure and, more importantly, its reversal in humans.

We hypothesize that a feedback regulatory mechanism(s) critical to blood formation is defective in scat. To identify mechanisms underlying the cyclic scat phenotype, we are performing high throughput sequencing (RNAseq) and global transcriptome analysis using tissues and purified cell populations from the scat disease stages (e.g., crisis and remission). Comprehensive transcriptome analysis will generate gene- and transcript-level expression values, define pathways/networks over-represented in differentially expressed gene lists, and identify functional gene modules correlated with disease status using powerful unbiased weighted gene co-expression network analysis (WGCNA). These analyses will define gene networks highly correlated with each progressive disease stage and uncover candidate “hub” genes for subsequent prioritization and functional testing as critical regulators of vertebrate hematopoiesis.

Anemia of aging

Anemia in aged (defined as >65 years of age) individuals is a common condition, affecting more than 3 million people (approximately 10%) of the elderly population in the United States. Indeed, the anemia of aging was recognized as so pervasive that the American Society of Hematology declared it a public health crisis in 2009. Epidemiology studies have consistently shown that anemia is an independent risk factor for disability and mortality in the elderly population. Even mild anemia is associated with enormous medical costs due to the large number of significant sequelae including a higher incidence of severe cardiovascular disease, cognitive impairment, poor outcomes from hospitalizations, and decreased physical function. The latter leads to difficulty performing everyday tasks, increased risk of falls and fractures, immobility, isolation, development of frailty and increased mortality in a high percentage of cases. Thus, anemia of aging has an enormous, negative impact on healthspan and quality of life not seen in younger people with anemia.

Our goal is to identify age-related factors influencing baseline hemoglobin levels and related erythroid traits such as the red blood cell count and hematocrit. To accomplish this goal, we have again turned to DO mice. DO mice closely mimic human genetic variation and enable us to obtain QTL with extremely high resolution. In conjunction with The Jackson Laboratory Nathan Shock Center for Excellence in the Basic Biology of Aging, we obtained, aged and phenotyped (complete blood counts) a longitudinal cohort of 600 DO mice at young (6 months of age), middle and old (18 months) ages. These mice are being densely genotyped using SNP arrays, and will be analyzed for QTL influencing hemoglobin levels and other related erythroid traits as a function of age. Ultimately, candidate genes influencing age-dependent changes in red cell parameters will be identified and validated.