Despite the importance of keratinocytes in epidermal structure and function, the molecular mechanisms regulating the division of these cells remain poorly understood. Herein, we demonstrate an essential role for cilia and flagella associated protein 100 (CFAP100) in keratinocyte division. Cfap100-knockout mice display a thinner and transparent skin and an impaired epidermal barrier function. Depletion of CFAP100 in keratinocytes prolongs mitotic progression and compromises chromosome segregation. Molecular studies reveal that CFAP100 interacts with tubulin tyrosine ligase-like protein 13 (TTLL13) to maintain spindle pole integrity in dividing keratinocytes. Further analysis shows that CFAP100 recruits TTLL13 to the spindle pole to increase the glutamylation of spindle microtubules. Restoring microtubule glutamylation by overexpression of TTLL13 or depletion of cytosolic carboxypeptidase 5 remarkably rescues the spindle pole defects in CFAP100-depleted cells. These findings thus identify CFAP100 as a central link to couple microtubule glutamylation to spindle pole integrity in keratinocytes to promote epidermal development.