Smokers may use nicotine to self-medicate for situation-specific or person-specific cognitive or affective deficits. Although evidence suggests that nicotine replacement therapy (NRT), relative to placebo, enhances spatial working memory (SWM) in smoking-abstinent smokers with schizophrenia, the extent to which NRT may be helpful in attenuating abstinence-related SWM in other groups with deficits in SWM is unknown. Depressive symptoms are associated with both tobacco smoking and deficits in SWM. Previous studies have found that smoking abstinence increases depressive affect and depression-related hemispheric asymmetries in brain activation. Although the serotonin neurotransmitter system is closely associated with depression and the effects of nicotine, the authors are not aware of any studies that have evaluated the possible role of individual differences in serotonin transporter (5-HTT) genotype and depressive symptoms as moderators of the effects of NRT on SWM. Thus, the current study assessed the effects of NRT (nicotine patch) on SWM in relation to: (1) depressive traits and (2) 5-HTT genotype. Smoking-deprived habitual smokers (N = 64) completed the dot recall test of SWM during counterbalanced and double-blind nicotine and placebo testing sessions. There was a marginal overall effect of NRT on SWM. More importantly, NRT enhanced SWM in 5-HTT short allele carriers, relative to those with two long alleles, and this enhancement in short-allele carriers was greater for individuals with higher levels of depressive symptoms.