Hoxa cluster genes determine the proliferative activity of adult mouse hematopoietic stem and progenitor cells

Determination of defined roles for endogenous homeobox (Hox) genes in adult hematopoietic stem and progenitor cell (HSPC) activity has been hampered by a combination of embryonic defects and functional redundancy. Here we show that conditional homozygous deletion of the Hoxa cluster (Hoxa−/−) results in a marked reduction of adult HSPC activity, both in vitro and in vivo. Specifically, proliferation of Hoxa−/− HSPCs is reduced compared with wild-type (WT) cells in vitro and they are less competitive in vivo. Notably, the loss of Hoxa genes had little impact on HSPC differentiation. Comparative RNA sequencing analyses of Hoxa−/− and WT hematopoietic stem cells (CD150+/CD48−/Lineage−/c-kit+/Sca-1+) identified a large number of differentially expressed genes, three of which (Nr4a3, Col1a1, and Hnf4a) showed >10-fold reduced levels. Engineered overexpression of Hoxa9 in Hoxa−/− HSPCs resulted in partial phenotypic rescue in vivo with associated recovery in expression of a large proportion of deregulated genes. Together, these results provide definitive evidence linking Hoxa gene expression to proliferation of adult HSPCs

Hoxa9 collaborates with E2A-PBX1 in mouse B cell leukemia in association with Flt3 activation and decrease of B cell gene expression

Background: The fusion protein E2A-PBX1 induces pediatric B cell leukemia in human. Previously, we reported oncogenic interactions between homeobox (Hox) genes and E2A-PBX1 in murine T cell leukemia. A proviral insertional mutagenesis screen with our E2A-PBX1 B cell leukemia mouse model identified Hoxa genes as potential collaborators to E2A-PBX1. Here we studied whether Hoxa9 could enhance E2A-PBX1 leukemogenesis. Results: We show that Hoxa9 confers a proliferative advantage to E2A-PBX1 B cells. Transplantation experiments with E2A-PBX1 transgenic B cells overexpressing Hoxa9 isolated from bone marrow chimeras showed that Hoxa9 accelerates the generation of E2A-PBX1 B cell leukemia, but Hoxa9 is unable to transform B cells alone. Quantitative-reverse transcriptase polymerase chain reaction analysis demonstrated a strong repression of B cell specific genes in these E2A-PBX1/Hoxa9 leukemias in addition to Flt3 activation, indicating inhibition of B cell differentiation in combination with enhanced proliferation. Overexpression of Hoxa9 in established E2A-PBX1 mouse leukemic B cells resulted in a growth advantage in vitro, which was also characterized by an enhanced expression of Flt3. Conclusions: we show for the first time that Hoxa9 collaborates with E2A-PBX1 in the oncogenic transformation of B cells in a mouse model that involves Flt3 signaling, which is potentially relevant to human disease

Transancestral mapping and genetic load in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease with marked gender and ethnic disparities. We report a large transancestral association study of SLE using Immunochip genotype data from 27,574 individuals of European (EA), African (AA) and Hispanic Amerindian (HA) ancestry. We identify 58 distinct non-HLA regions in EA, 9 in AA and 16 in HA (B50% of these regions have multiple independent associations); these include 24 novel SLE regions (Po5 10 8), refined association signals in established regions, extended associations to additional ancestries, and a disentangled complex HLA multigenic effect. The risk allele count (genetic load) exhibits an accelerating pattern of SLE risk, leading us to posit a cumulative hit hypothesis for autoimmune disease. Comparing results across the three ancestries identifies both ancestry-dependent and ancestry-independent contributions to SLE risk. Our results are consistent with the unique and complex histories of the populations sampled, and collectively help clarify the genetic architecture and ethnic disparities in SL

Hoxb4 overexpression in CD4 memory phenotype T cells increases the central memory population upon homeostatic proliferation.

Memory T cell populations allow a rapid immune response to pathogens that have been previously encountered and thus form the basis of success in vaccinations. However, the molecular pathways underlying the development and maintenance of these cells are only starting to be unveiled. Memory T cells have the capacity to self renew as do hematopoietic stem cells, and overlapping gene expression profiles suggested that these cells might use the same self-renewal pathways. The transcription factor Hoxb4 has been shown to promote self-renewal divisions of hematopoietic stem cells resulting in an expansion of these cells. In this study we investigated whether overexpression of Hoxb4 could provide an advantage to CD4 memory phenotype T cells in engrafting the niche of T cell deficient mice following adoptive transfer. Competitive transplantation experiments demonstrated that CD4 memory phenotype T cells derived from mice transgenic for Hoxb4 contributed overall less to the repopulation of the lymphoid organs than wild type CD4 memory phenotype T cells after two months. These proportions were relatively maintained following serial transplantation in secondary and tertiary mice. Interestingly, a significantly higher percentage of the Hoxb4 CD4 memory phenotype T cell population expressed the CD62L and Ly6C surface markers, characteristic for central memory T cells, after homeostatic proliferation. Thus Hoxb4 favours the maintenance and increase of the CD4 central memory phenotype T cell population. These cells are more stem cell like and might eventually lead to an advantage of Hoxb4 T cells after subjecting the cells to additional rounds of proliferation

Medium-term competitive homeostatic proliferations (60 days) of <i>Hoxb4</i> transgenic and wt CD4 MP T cells.

<p>(<b>A</b>) FACS profile showing fractions of <i>Hoxb4</i> and wt cells to donor derived MP T population in lymph node (LN). (<b>B</b>) Stacked bar graphs indicating the average contributions of <i>Hoxb4</i> and wt cells in LN, Spl and BM measured in three independent experiments; n = 9. (<b>C</b>) FACS profiles for the expression of typical memory T cell surface markers on <i>Hoxb4</i> and wt MP T cells in the BM. (<b>D</b>) Average subpopulations of <i>Hoxb4</i> and wt MP T cells expressing the indicated surface markers in LN (upper panel), Spl (middle panel) and BM (lower panel). (<b>E</b>) Percentage of <i>Hoxb4</i> and wt MP T cells (gated on CD44^hi) positive for indicated cytokines (n = 3–6). *P<0.05, 2-tailed Student ttest. MP = memory phenotype, wt = wild type, LN = lymph node, Spl = spleen and BM = bone marrow, TNF = tumor necrosis factor; IL-2 = interleukine-2; IFN = interferon.</p

Long-term competitive homeostatic proliferations (180 days) of <i>Hoxb4</i> transgenic and wt CD4 cells.

<p>(<b>A</b>) Scheme of serial transplantations. 10×10⁶ cells of the LNs of primary hosts that received a transplant composed of equal doses of <i>Hoxb4</i> and wt MP T cells were serially transplanted into secondary and tertiary hosts with a 60 days interval. (<b>B</b>) Compilation of <i>Hoxb4</i> and wt fractions of donor derived cells in LN, Spl and BM of secondary (n = 6) and tertiary hosts (n = 4) from two independent experiments. (<b>C</b>) Bar graphs showing the average percentage of cells positive for CD62L and Ly6C within the <i>Hoxb4</i> or wt memory T populations. *P<0.05, 2-tailed Student ttest. Wt = wild type, MP = memory phenotype, LN = lymph node, Spl = spleen and BM = bone marrow.</p

Change of naïve and MP T cell populations in <i>Hoxb4</i> transgenic and wt mice with age.

<p>Scatter plots showing the percentage of (<b>A</b>) CD4 and (<b>B</b>) CD8 T cells that are naïve (CD44^lo/CD62L^hi), MP (CD44^hi) or are a subpopulation of MP T cells (CD44^hi/Ly6C^hi) for LN, Spl and BM derived from individual <i>Hoxb4</i> transgenic and wt (n = 6–8) age matched mice. Young mice are between 3–4 months and old mice are all older than 28 months. Naïve and MP populations change significantly with age, but not between <i>Hoxb4</i> and wt mice. *P<0.05, 2-tailed Student ttest. MP = memory phenotype, Wt = wild type, LN = lymph node, Spl = spleen and BM = bone marrow.</p

Total donor cells contribution to hematopoietic organs and the proportion of <i>Hoxb4</i> versus wild type cells.

<p>LN = Lymph node; BM = bone marrow, wt = wild type.</p