Chemoresistance is a significant challenge and major obstacle to achieving cancer remission during chemotherapy, primarily due to the risk of recurrence and metastasis. This study reveals that linker histone H1.0 plays a crucial role in paclitaxel resistance (TXR) in ovarian cancer cells by regulating Histone deacetylase 5 (HDAC5), which deacetylates core histones and represses gene transactivation. Transcriptomic profile analysis revealed that cytokine signaling networks are enriched pathways that correlate with H1.0 expression. Advanced clustering analysis identified interleukin 6 (IL-6) as a key molecule connecting these enriched H1.0-related pathways. Furthermore, gain- and loss-of-H1.0 expression experiments showed that H1.0 controls IL-6 mRNA and protein expression in ovarian cancer cells. Additionally, our findings indicate that HDAC5 expression is downregulated in SKOV3/Txr cells compared with parental cells. H1.0 silencing in TXR cells increases HDAC5 levels, suggesting an antagonistic effect between H1.0 and HDAC5. Cell viability assays showed that HDAC5 overexpression markedly inhibited cell survival. Furthermore, ectopic HDAC5 overexpression reduced IL-6 mRNA and protein expression, which was increased by H1.0. This effect was associated with reduced H3K9Ac core histone acetylation and decreased NF-κB binding on the IL-6 promoter, as demonstrated by chromatin immunoprecipitation assays. Further analysis revealed that HDAC5 is downregulated in several tumor types. Furthermore, high H1.0 and IL-6 expression, coupled with low HDAC5 levels, was exclusively observed in ovarian carcinoma. Together, our results demonstrate an interplay between H1.0, HDAC5, and IL-6 in modulating paclitaxel resistance in ovarian cancer cells, highlighting new therapeutic targets to overcome chemoresistance.