"Maesetsu! Opening Act" centers on four aspiring comedians navigating the challenges of breaking into the competitive world of manzai, a traditional Japanese comedic performance style involving a straight man (tsukkomi) and a funny man (boke). The story primarily follows two pairs of childhood friends: Mafuyu Kogarashi and Fubuki Kitakaze, and Rin Araya and Nayuta Asogi. Both duos share a passion for comedy but face different struggles as they strive to make their mark in the industry.

Mafuyu and Fubuki, known as "Fastener," are a high-energy duo with contrasting personalities. Mafuyu, the tsukkomi, is serious and hardworking, while Fubuki, the boke, is carefree and often relies on her natural charm. Their dynamic is tested as they balance their comedic ambitions with the realities of daily life, including part-time jobs and personal insecurities. Meanwhile, Rin and Nayuta, performing as "Rinayata," are a more reserved pair. Rin, the tsukkomi, is methodical and detail-oriented, while Nayuta, the boke, is quirky and unpredictable. Their partnership is built on mutual respect, but they struggle to find their unique voice in a crowded field.

The series explores the characters' growth as they participate in local comedy competitions, attend workshops, and interact with established comedians. Each episode highlights their efforts to refine their craft, often blending humor with moments of introspection. The narrative also delves into their relationships with family, friends, and mentors, providing a grounded perspective on their journey. Themes of perseverance, friendship, and self-discovery are woven throughout, as the characters learn to overcome setbacks and embrace their individuality.

Supporting characters, such as their peers and industry professionals, add depth to the story by offering guidance, competition, and occasional comic relief. The series balances lighthearted moments with the realities of pursuing a creative career, portraying both the joys and frustrations of chasing a dream. Through its focus on the characters' personal and professional lives, "Maesetsu! Opening Act" offers a heartfelt and relatable portrayal of young comedians striving to find their place in the world of manzai.