Objective In the present study, we assessed the associations among fatigue, quality of life (QOL), clinical parameters, and body mass index (BMI) with autonomic function in end-stage renal disease (ESRD) patients undergoing hemodialysis as well as fatigue-free healthy subjects. Design and Methods This was a case-control study. This study compared autonomic function in ESRD patients (n = 192) to that of healthy subjects (n = 282) and evaluated its association with fatigue, QOL, and clinical parameters such as glucose, albumin, cholesterol, and BMI. Fatigue was evaluated by a recently established fatigue questionnaire and performance status, and QOL was evaluated with the kidney disease QOL questionnaire. With regards to autonomic function, spontaneous beat-to-beat variations were measured, according to time- (standard deviation of all normal a-wave intervals [CVa-a%]) and frequency domains (low frequency [LF] power, high frequency [HF] power, and LF/HF ratio) with acceleration plethysmography. Results CVa-a%, LF power, HF power, and LF/HF ratio were significantly lower in ESRD patients than healthy subjects. There were significant inverse correlations between these factors and age in healthy subjects, but not in ESRD patients. Although the fatigue score was not associated with any autonomic parameters, ESRD patients with impaired performance status exhibited a significantly lower LF/HF ratio. Moreover, in ESRD patients, the LF/HF ratio was significantly and positively associated with several components of QOL, including physical functioning and role emotional, independent of other clinical parameters and BMI. Conclusions Impaired autonomic function is significantly associated with fatigue and impaired QOL in dialysis patients.