Neurodegenerative disorders, such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), are increasing in prevalence but lack targeted and mechanism-based therapies. Despite various biological stresses and processes that appear to contribute to the pathogenesis of these diseases, the molecular events and converging pathways remain unclear. We recently identified mutations in the CCNF gene as a novel cause of ALS/FTD, with the Ser621Gly mutation found to segregate across multiple generations in an Australian family. CCNF encodes cyclin F, an E3 ubiquitin ligase that forms a part of a SCF complex that binds to proteins for ubiquitylation and degradation by the ubiquitin-proteasome system (UPS). Experimental expression of the cyclin FS621G mutation led to defective protein degradation, motor axonopathy, and features of ALS pathogenesis in vitro and in vivo. We investigated the effect of the cyclin FS621G mutation on Lys48-specific ubiquitylation, and how this mutation alters its E3 ligase activity and stability that contributes to the ubiquitylation of neuronal proteins and causes proteostasis dysfunction. Additionally, we examined the phosphorylation status of cyclin F at Ser621 and how this site regulates the Lys48-specific ubiquitylation activity of the SCF(Cyclin F) complex. Proteomic analysis of immunoprecipitated Lys48-ubiquitylated proteins from mutant cyclin FS621G identified proteins that clustered to the autophagy pathway, including sequestosome-1 (p62/SQSTM1), heat shock proteins (HSPs) and chaperonin complex components. Examination of autophagy markers p62, LC3 and Lamp2 in mutant cyclin FS621G revealed autophagy defects specifically resulting in impairment in autophagosomal-lysosome fusion. We also identified a mechanism by which cyclin F hyperubiquitylates p62, the receptor responsible for transporting substrates for autophagic degradation. These findings demonstrate that a single missense mutation in ALS/FTD-causing cyclin F disrupts Lys48-specific ubiquitylation, leading to accumulation of substrates and defects to autophagy, that are pathological events leading up and/or contribute to ALS and FTD progression.