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The cytoskeletal system of a cell in embryonic development is critical for the asymmetric distribution of transmembrane and cytosolic proteins of the PCP pathway. Cytoskeletal regulation, specifically the regulation of non-centrosomal MT, enable proximal-distal (P-D) transportation of proteins and effectors within a cell. . Moreover, MTs also provide the mechanical support necessary to support PCP.

The molecular mechanism that specifies the global orientation of the P-D ends of cells within a tissue remains elusive. Many papers however, suggest that this is regulated by the atypical cadherins Ds and Ft; and the golgi kinase Fj. Initially, Fj is differentially expressed across a tissue, with higher distal expression and lower proximal expression. During post-translational modification (PTM), Fj uses its kinase domain to phosphorylate three cadherin domains of Ds ; this reduces Ds' binding affinity with its ligand, Ft. Conversely, Fj's PTM of Ft increases its binding affinity with Ds. The result of Fj's differential effects on these atypical cadherins, coupled with its differential expression along the P-D axis, is the creation of opposing gradients of Ds and Ft; Ds is concentrated proximally and Ft is concentrated distally. As transmembrane proteins, Ds and Ft form heterodimeric interactions between adjacent cells. This interaction plays a role in orientating the plus end of the apical non-centrosomal MT towards the distal side of the cell. Although the exact molecular mechanism remains cryptic, studies have shown that as the angle of MTs approach the P-D axis, the stability of tubulin and the rate of polymerisation increases. This process has been implicated in Drosophila, Xenopus and Mus musculus. For example, Mus musculus possess homologues for Fz (Fzd3, Fzd6), Stbm (Vangl2), Fmi (Celsr1), Dsh (Dvl1, Dvl2), Ft (Fat4) and Ds (Dchs1 and Dchs2). These proteins have been implicated in interactions similar to the pathways described in the Drosophila. This suggests that these proteins place a high pressure on natural selection to conserve them.

The organisation of MT by the Ft, Ds, Fj system has been observed in the Drosophila wing and posterior abdomen. Surprisingly, the opposite behaviour is observed in Drosophila eyes and anterior abdomen, where the plus end of the non-centrosomal MT is orientated proximally. It's been suggested that this opposite behaviour is a function of the relative abundance of the alternatively-spliced Pk isoforms, Pricklepk and Pricklespiny-legs. Overexpression studies found that the over-abundance of Pricklepk in early embryogenesis leads to MT nucleation at the proximal side of a drosophila wing cell, then plus end polymerisation towards the distal membrane. Consequently, this ensures the distal transport of vesicles containing Fz, Dsh and Fmi. Conversely, the over-abundance of Pricklespiny-legs leads to distal MT nucleation and proximal MT polymerisation, which leads to Fz, Dsh and Fmi vesicle transportation towards the opposite direction. This result is consistent with the observation of increased Pricklespiny-legs expression in the WT Drosophila eyes and anterior abdomen. The MT directional bias within a Drosophila wing, caused by the Pk isoforms is dependent on the Fs, Ds, Fj system; this suggests that the Pk isoforms determine how MTs interpret the Ds-Fj gradient, hence their biased orientation. Similarly, the roles for the two Pk isoforms within the axon of a neuron has also been explored. Researchers found that a higher abundance of Pricklepk promotes plus-ended growth of MTs and vesicle transport towards the synapse, while a higher abundance of Pricklespiny-legs promotes these activities towards the cell body.

Although molecular asymmetry plays a key role in PCP, the mechanical morphogenesis of cells is another aspect of PCP that remains to be fully understood. A study on the Drosophila wing found that MTs and myosin II (MyoII) create opposing forces along the P-D axis at the adherens junctions (AJ) level. It should be noted that MTs are dynamic structures that are constantly going through polymerisation and depolymerisation. As MTs reach the cortex of the cell, it exerts polarised mechanical pressure upon the membrane which morphs the cell shape; MyoII acts as a counterbalance to prevent the cell from breaking apart. This association creates a tissue-level structural support, which enables the stability of polarised, trans-cellular structure along the P-D axis.