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With-No-Lysine (WNK) kinases are a subfamily of serine/threonine protein kinases. WNKs are involved in plant abiotic stress response and circadian rhythms. However, members of the WNK subfamily and their responses to abiotic and biotic stresses in <i>Gossypium hirsutum</i> have not been reported. In this study, 26 <i>GhWNKs</i> were identified in <i>G. hirsutum</i>. The gene structure, conserved motifs, and upstream open reading frames (uORFs) of <i>GhWNKs</i> were identified. Moreover, <i>GhWNKs</i> regulation is predicted to be regulated by <i>cis</i>-acting elements, such as ABA responsive element (ABRE), MBS, and MYC. Furthermore, transcription factors including MIKC_MADS, C2H2, TALE, bZIP, Dof, MYB, bHLH, and HD-ZIP are projected to play a regulatory role in <i>GhWNKs</i>. The expression patterns of <i>GhWNKs</i> under normal conditions and biotic and abiotic stresses were evaluated, and their expression was found to vary. The expression patterns of several <i>GhWNKs</i> were induced by infiltration with <i>Verticillium dahliae</i>, suggesting that several <i>GhWNKs</i> may play important roles in the response of cotton to <i>V. dahliae</i>. Interestingly, a homoeologous expression bias within the <i>GhWNKs</i> was uncovered in upland cotton. Homoeologous expression bias within <i>GhWNKs</i> provides a framework to assist researchers and breeders in developing strategies to improve cotton traits by manipulating individual or multiple homeologs.