Acquirement of resistance to rituximab has been observed in lymphoma patients. To define mechanisms associated with rituximab resistance, we developed various rituximab-resistant cell lines (RRCL) and studied changes in CD20 expression/structure, lipid raft domain (LRD) reorganization, calcium mobilization, antibody-dependent cellular cytotoxicity, and complementmediated cytotoxicity (CMC) between parental and RRCL. Significant changes in surface CD20 antigen expression were shown in RRCL. Decreased calcium mobilization and redistribution of CD20 into LRD were found in RRCL. Western blotting identified a unique 35 kDa protein band in RRCL, which was not seen in parental cells and was secondary to an increase in surface and cytoplasmic expression of IgM light chains. CD20 gene expression was decreased in RRCL. In vitro exposure to PS341 increased CD20 expression in RRCL and minimally improved the sensitivity to rituximab-associated CMC. Our data strongly suggest that the acquisition of rituximab resistance is associated with global gene and protein down-regulation of the CD20 antigen affecting LRD organization and downstream signaling. CD20 expression seems to be regulated at the pretranscriptional and posttranscriptional levels. Proteasome inhibition partially reversed rituximab resistance, suggesting the existence of additional mediators of rituximab resistance. Future research is geared to identify drugs and/or biological agents that are effective against RRCL.

Recently, a unique form of X chromosome dosage compensation has been demonstrated in human preimplantation embryos, which happens through the dampening of X-linked gene expression from both X chromosomes. Subsequently, X chromosome dampening has also been demonstrated in female human pluripotent stem cells (hPSCs) during the transition from primed to naive state. However, the existence of dampened X chromosomes in both embryos and hPSCs remains controversial. Specifically, in preimplantation embryos it has been shown that there is inactivation of X chromosome instead of dampening. Here, we performed allelic analysis of X-linked genes at the single-cell level in hPSCs and found that there is partial reactivation of the inactive X chromosome instead of chromosome-wide dampening upon conversion from primed to naive state. In addition, our analysis suggests that the reduced X-linked gene expression in naive hPSCs might be the consequence of erasure of active X chromosome upregulation.

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