Reticular Dysgenesis-Associated Adenylate Kinase 2 Deficiency Has Opposing Metabolic Consequences on Granulopoiesis and Lymphopoiesis
Authors: Wang Wenqing , Arreola Martin , Mohammed Abdulvasey , Weinacht Katja G. 
Published: December 2024

Metabolic disorders impacting hematopoiesis have provided unique insights into the regulatory networks that maintain metabolic homeostasis during differentiation of hematopoietic stem and progenitor cells (HSPCs). Reticular Dysgenesis (RD), a particularly grave form of severe combined immunodeficiency (SCID), is caused by biallelic loss of function mutations in mitochondrial adenylate kinase 2 (AK2). The hematopoietic phenotype of RD manifests in defective myeloid and lymphoid development, resulting in profound neutropenia and lymphopenia. AK2 catalyzes the reversible phosphorylation of adenosine monophosphate (AMP) to adenosine diphosphate (ADP) in the mitochondrial intermembrane space. ADP is used to fuel adenosine triphosphate (ATP) production. While there is an abundance of processes that produce ADP, AK2 contributes to the vast majority of AMP “recycling” activities in the myeloid and lymphoid lineages. We have previously shown that AK2 deficiency leads to a steep increase in cellular AMP/ATP and AMP/ADP ratios, but how this leads to the demise of granulopoiesis and lymphopoiesis in RD patients remains unclear.

To assess the mechanistic basis of RD, we performed single-cell RNA-sequencing on bone marrow samples of two previously reported patients with biallelic AK2 c.542G&gt;A, p.R175Q missense mutations, and nine healthy donor controls. A gene set enrichment analysis revealed that AK2 deficiency had opposing effects on the granulocytic and lymphoid lineages. While anabolic pathways related to ribonucleoprotein synthesis were paradoxically upregulated in granulocytes from RD patients, the same pathways were profoundly downregulated in lymphoid cells from RD patients relative to healthy donors.

To further investigate how different hematopoietic lineages respond to metabolic stress caused by AK2 deficiency, we have developed an inducible Ak2 knockout mouse model under the control of Mx1-Cre that specifically targets hematopoietic cells (Ak2fl/fl; Mx1-Cre). We found that the consequences of Ak2 deficiency for the hematopoietic system are contingent on the effective engagement of metabolic checkpoints. In hematopoietic stem and progenitor cells, Ak2 deficiency mildly reduced mechanistic target of rapamycin (mTOR) signaling and anabolic pathway activation. This conserved nutrient homeostasis and maintained cell survival. Later during granulopoiesis, metabolic checkpoints became ineffective, leading to a paradoxical upregulation of mTOR activity and energy-consuming anabolic pathways. This caused nucleotide imbalance and the depletion of essential substrates, ultimately resulting in proliferation arrest and demise of the granulocyte lineage. In developing T cells (thymocytes), however, mTOR activity and anabolic pathways remain profoundly suppressed, and Ak2-deficient thymocytes failed to mature past the double-positive (DP) stage. Notably, T cell receptor (TCR) positive cells were completely absent in Ak2-deficient thymocytes, suggesting that Ak2 activity is indispensable for TCR rearrangement. Our findings suggest that although intricate metabolic checkpoint control may help tolerate severe metabolic defects, the failure or overactivity of these checkpoints, as observed in granulocytic and T lineages, both result in a catastrophic loss of metabolic homeostasis and failure of maturation. Our ongoing work characterizes the essential role of AK2 during intra-thymic T cell development.