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Editorials
Genetic fingerprint defines hematopoietic stem cell pool size and function
Tatsuya Morishima1,2 and Hitoshi Takizawa1,3
1Laboratory of Stem Cell Stress, International Research Center for Medical Sciences, Kumamoto University; 2Laboratory of Hematopoietic Stem Cell Engineering, International Research Center for Medical Sciences, Kumamoto University and 3Center for Metabolic Regulation of Healthy Aging, Kumamoto University, Kumamoto, Japan
E-mail: HITOSHI TAKIZAWA - htakizawa@kumamoto-u.ac.jp doi:10.3324/haematol.2019.241299
Hematopoietic stem cells (HSC) are at the apex of the hematopoietic tree, with self-renewal and multilineage differentiation potential. On the one hand, HSC can replenish the mature blood cells by differ- entiating into lineage-committed progenitor cells in response to the shortage of blood cells under both home- ostatic and stressed conditions, such as bleeding and infection. On the other hand, HSC replicate themselves to maintain their number. This differentiation and self- renewal needs to be strictly regulated by gene expression regulation in order to maintain life-long hematopoiesis.1 Gene expression is in general regulated by “cis- and trans- regulatory elements”.2 Trans-regulatory elements are defined as factors which regulate expression of distal genes (e.g. transcription factors), while cis-regulatory ele- ments are defined as non-coding DNA sequences which regulate expression of proximal genes (e.g. promoter and enhancer regions). These cis-regulatory elements play important roles in evolution in which polymorphisms occur in cis-regulatory elements and contribute to pheno- typic diversity of organisms within well-conserved genes.3 Epigenetics plays important roles in HSC regula- tion, as epigenetic dysregulation in HSC is a key driver for HSC aging and hematopoietic malignancies.4 Epigenetic regulation is also controlled by cis- and trans- regulatory elements. For example, histone modifications function as trans-regulatory elements, whereas DNA methylations function as cis-regulatory elements.
Phenotypic diversity is frequently caused by genetic variations such as single nucleotide polymorphism (SNP). It was reported that the size and function of the HSC pool vary between mice strains,5-9 which suggests genetic background, such as SNP and copy number variations, define HSC homeostasis. In 2007, Liang et al. identified latexin (Lxn) as an HSC regulatory gene whose expression level is inversely correlated with HSC number.10 Although a variation in Lxn gene expression and HSC number in different tested mouse strains was shown, the mecha- nism underlying regulation of Lxn gene expression and its variation between mice strains remained unknown.
In this issue of Haematologica, the same group who published the above-mentioned paper,10 Zhang et al. iden- tified the promoter region of the Lxn gene that controls the level of Lxn gene expression via both HMGB2, a chro- matin protein, and genetic variations in the promoter region.11 To study the transcriptional regulation of Lxn, the authors characterized the upstream region of the Lxn gene. Based on the natural variation of Lxn expression, they searched SNP with CpG island and identified a region with strong promoter activity in the upstream. Subsequently, DNA pulldown in combination with mass spectrometry analysis were performed to identify pro-
or TATSUYA MORISHIMA - tatsuyam@kumamoto-u.ac.jp
teins bound to this region, and HMGB2 was found to bind to the promoter region and to suppress Lxn gene promoter activity. To further confirm the regulatory role of HMGB2 in Lxn gene expression, Zhang et al. per- formed a gene knockdown experiment with EML cells, which share some of the characteristics of HSC.12 This showed that HMGB2 knockdown suppresses EML cell growth and that additional Lxn gene knockdown could rescue this growth, suggesting that Lxn was one of the transcriptional targets of HMGB2. Consistent with the previous reports concerning the phenotype of cells over- expressing Lxn,13,14 the HMGB2 knockdown cells showed enhanced apoptosis and cell cycle arrest, which could in part be rescued by Lxn gene knockdown. These data sug- gest HMGB2 positively regulates HSC survival and prolif- eration by suppressing expression of Lxn and Lxn-regulat- ed apoptosis. Similar data were also shown in Lin-Sca-1+c- Kit+ (LSK) cells primarily isolated from mouse bone mar- row that contain HSC. HMGB2 knockdown in LSK cells showed suppressed proliferation, enhanced apoptosis and cell cycle arrest in vitro. In transplantation experi- ments, HMGB2 knockdown in LSK cells showed decreased reconstitution of whole peripheral blood cells and bone marrow LSK cells and long-term HSC in trans- plant recipients, indicating that HMGB2 plays an impor- tant role in HSC function in vivo.
The previous finding that Lxn expression level is correlat- ed with HSC numbers10 led the authors to hypothesize that the SNP in the promoter region of the Lxn gene may contribute to a variation in Lxn expression and HSC num- ber. To test this, the authors introduced G to C mutation in the HMGB2 binding sequence in the Lxn gene promoter region and found that the G allele showed higher promoter activity for Lxn expression compared to the C allele. Furthermore, when Lxn gene expression and bone marrow HSC number were analyzed in different mouse strains car- rying the G or C allele in this SNP region, the mice strain carrying the C allele showed relatively lower Lxn protein expression and higher HSC number compared to those car- rying the G allele. These data suggest that a genetic variant in the Lxn gene promoter region defines the variation in Lxn gene expression level and HSC number.
Together, Zhang et al. revealed that the transcription of Lxn regulating HSC function, at least as far as apoptosis is concerned, was controlled by both trans-regulatory ele- ment, HMGB2, and cis-regulatory element, as genetic variation was observed in the Lxn gene promoter region (Figure 1). HMGB2 is known as a chromatin-associated protein which remodels chromatin structure and gene expression.15 Although the molecular mechanism by which HMGB2 regulates Lxn gene transcription remains unclear, all the data provided in this study suggest Lxn
haematologica | 2020; 105(3)