RESUMEN
T-cell acute lymphoblastic leukemia (T-ALL) is one of the deadliest and most aggressive hematological malignancies, but its pathological mechanism in controlling cell survival is not fully understood. Oculocerebrorenal syndrome of Lowe is a rare X-linked recessive disorder characterized by cataracts, intellectual disability, and proteinuria. This disease has been shown to be caused by mutation of oculocerebrorenal syndrome of Lowe 1 (OCRL1; OCRL), encoding a phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] 5-phosphatase involved in regulating membrane trafficking; however, its function in cancer cells is unclear. Here, we uncovered that OCRL1 is overexpressed in T-ALL cells, and knockdown of OCRL1 results in cell death, indicating the essential role of OCRL in controlling T-ALL cell survival. We show OCRL is primarily localized in the Golgi and can translocate to plasma membrane (PM) upon ligand stimulation. We found OCRL interacts with oxysterol-binding protein-related protein 4L, which facilitates OCRL translocation from the Golgi to the PM upon cluster of differentiation 3 stimulation. Thus, OCRL represses the activity of oxysterol-binding protein-related protein 4L to prevent excessive PI(4,5)P2 hydrolysis by phosphoinositide phospholipase C ß3 and uncontrolled Ca2+ release from the endoplasmic reticulum. We propose OCRL1 deletion leads to accumulation of PI(4,5)P2 in the PM, disrupting the normal Ca2+ oscillation pattern in the cytosol and leading to mitochondrial Ca2+ overloading, ultimately causing T-ALL cell mitochondrial dysfunction and cell death. These results highlight a critical role for OCRL in maintaining moderate PI(4,5)P2 availability in T-ALL cells. Our findings also raise the possibility of targeting OCRL1 to treat T-ALL disease.
Asunto(s)
Membrana Celular , Fosfatidilinositol 4,5-Difosfato , Monoéster Fosfórico Hidrolasas , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Linfocitos T , Humanos , Membrana Celular/metabolismo , Supervivencia Celular , Hidrólisis , Síndrome Oculocerebrorrenal/enzimología , Síndrome Oculocerebrorrenal/genética , Fosfatidilinositol 4,5-Difosfato/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/inmunología , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patología , Linfocitos T/citología , Linfocitos T/inmunología , Monoéster Fosfórico Hidrolasas/biosíntesis , Monoéster Fosfórico Hidrolasas/deficiencia , Monoéster Fosfórico Hidrolasas/genética , Monoéster Fosfórico Hidrolasas/metabolismo , Aparato de Golgi/metabolismo , Ligandos , Transporte de Proteínas , Señalización del Calcio , Mitocondrias/metabolismo , Mitocondrias/patología , Citosol/metabolismoRESUMEN
Leukemia stem cells (LSCs) are a rare subpopulation of abnormal hematopoietic stem cells (HSCs) that propagates leukemia and are responsible for the high frequency of relapse in therapies. Detailed insights into LSCs' survival will facilitate the identification of targets for therapeutic approaches. Here, we develop an inhibitor, LYZ-81, which targets ORP4L with high affinity and specificity and selectively eradicates LCSs in vitro and in vivo. ORP4L is expressed in LSCs but not in normal HSCs and is essential for LSC bioenergetics and survival. It extracts PIP2 from the plasma membrane and presents it to PLCß3, enabling IP3 generation and subsequent Ca2+-dependent bioenergetics. LYZ-81 binds ORP4L competitively with PIP2 and blocks PIP2 hydrolysis, resulting in defective Ca2+ signaling. The results provide evidence that LSCs can be eradicated through the inhibition of ORP4L by LYZ-81, which may serve as a starting point of drug development for the elimination of LSCs to eventually cure leukemia.
Asunto(s)
Células Madre Hematopoyéticas/efectos de los fármacos , Leucemia/metabolismo , Células Madre Neoplásicas/efectos de los fármacos , Fosfatidilinositol 4,5-Difosfato/metabolismo , Receptores de Esteroides/metabolismo , Animales , Antineoplásicos/farmacología , Línea Celular Tumoral , Membrana Celular/metabolismo , Células Madre Hematopoyéticas/metabolismo , Humanos , Leucemia/sangre , Ratones , Ratones Endogámicos NOD , Ratones SCID , Células Madre Neoplásicas/metabolismo , Fosfolipasa C beta/metabolismo , Receptores de Esteroides/antagonistas & inhibidoresRESUMEN
Phosphoinositide phospholipases C (PLCs) are a family of eukaryotic intracellular enzymes with important roles in signal transduction. In addition to their location at the plasma membrane, PLCs also exist within the cell nucleus where they are stored. We previously demonstrated that OSBP-related protein 4L (ORP4L) anchors cluster of differentiation 3ϵ (CD3ϵ) to the heterotrimeric G protein subunit (Gαq/11) to control PLCß3 relocation and activation. However, the underlying mechanism by which ORP4L facilitates PLCß3 translocation remains unknown. Here, using confocal immunofluorescence microscopy and coimmunoprecipitation assays, we report that ORP4L stimulates PLCß3 translocation from the nucleus to the plasma membrane in Jurkat T-cells in two steps. First, we found that ORP4L is required for the activation of Ras-related nuclear protein (RAN), a GTP-binding nuclear protein that binds to exportin 1 and eventually promotes the nuclear export of PLCß3. Second, we also observed that ORP4L interacts with vesicle-associated membrane protein-associated protein A (VAPA) through its two phenylalanines in an acidic tract (FFAT) motif. This complex enabled PLCß3 movement to the plasma membrane, indicating that PLCß3 translocation occurs in a VAPA-dependent manner. This study reveals detailed mechanistic insight into the role of ORP4L in PLCß3 redistribution from storage within the nucleus to the plasma membrane via RAN activation and interaction with VAPA in Jurkat T-cells.