A slab-column joint in a flat plate structure may exhibit upward or downward punching shear (UPS and DPS) failure when subjected to different abnormal loading conditions. This can cause distinct residual deformations of the slabs, which in turn influences the behavior of the consequent progressive collapse of the entire structure. This paper presents two experimental tests on two 2×2-bay, 1/3-scaled flat plate substructure specimens. Both tests were conducted in two loading phases. In the first loading phase (LP-1), the interior column of specimens UPS-Sub and DPS-Sub was subjected to a gradually increased upward and downward concentrated load, respectively, until punching failure occurred at the interior slab-column joint. In the second loading phase (LP-2), an incremented downward uniformly distributed load was applied on the slab of both specimens up until the ultimate state just before a complete collapse of the slab. After LP-1, local and global damage patterns were observed respectively from UPS-Sub and DPS-Sub. The punching shear strength and the corresponding displacement of the interior joint in UPS-Sub were found to be 58.2% higher and 53.6% lower, respectively, than the corresponding ones in DPS-Sub. In LP-2, due to the upward residual deformation of the slab in UPS-Sub as a result of LP-1, its first peak load, vertical displacement, and horizontal displacement under compressive membrane action were, respectively, 8.4%, 91.1%, and 57.1% larger than those in DPS-Sub. On the other hand, the postpunching peak load of the two specimens under tensile membrane action were identical. To offer an in-depth understanding of the mechanical behavior that resulted from upward or download punching shear failure, the punching shear strengths of the interior slab-column joints and collapse resistance of the substructures were also analytically investigated.